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<div class="quarto-title">
<h1 class="title">CPTR9_DE_Analysis</h1>
</div>
<div class="quarto-title-meta">
</div>
</header>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb1"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb1-1"><a href="#cb1-1" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(GeomxTools)</span>
<span id="cb1-2"><a href="#cb1-2" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(dplyr)</span>
<span id="cb1-3"><a href="#cb1-3" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(limma)</span>
<span id="cb1-4"><a href="#cb1-4" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(edgeR)</span>
<span id="cb1-5"><a href="#cb1-5" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(ggplot2)</span>
<span id="cb1-6"><a href="#cb1-6" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(ggrepel)</span>
<span id="cb1-7"><a href="#cb1-7" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(stringr)</span>
<span id="cb1-8"><a href="#cb1-8" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(PCAtools)</span>
<span id="cb1-9"><a href="#cb1-9" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(readxl)</span>
<span id="cb1-10"><a href="#cb1-10" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(gridExtra)</span>
<span id="cb1-11"><a href="#cb1-11" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(grid)</span>
<span id="cb1-12"><a href="#cb1-12" aria-hidden="true" tabindex="-1"></a><span class="fu">library</span>(DSPWorkflow)</span>
<span id="cb1-13"><a href="#cb1-13" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb1-14"><a href="#cb1-14" aria-hidden="true" tabindex="-1"></a><span class="co"># Source the helper functions</span></span>
<span id="cb1-15"><a href="#cb1-15" aria-hidden="true" tabindex="-1"></a><span class="fu">source</span>(<span class="st">"DSP_functions.R"</span>)</span>
<span id="cb1-16"><a href="#cb1-16" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb1-17"><a href="#cb1-17" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb1-18"><a href="#cb1-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb1-19"><a href="#cb1-19" aria-hidden="true" tabindex="-1"></a><span class="co"># Results folder where the results should be exported</span></span>
<span id="cb1-20"><a href="#cb1-20" aria-hidden="true" tabindex="-1"></a>results.folder <span class="ot"><-</span> <span class="st">"results/"</span></span>
<span id="cb1-21"><a href="#cb1-21" aria-hidden="true" tabindex="-1"></a>run.folder <span class="ot"><-</span> <span class="st">"7_23_2024/"</span></span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
</div>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb2"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb2-1"><a href="#cb2-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Load the R object containing normalized reads</span></span>
<span id="cb2-2"><a href="#cb2-2" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb2-3"><a href="#cb2-3" aria-hidden="true" tabindex="-1"></a>normalized.object <span class="ot"><-</span> <span class="fu">readRDS</span>(<span class="st">"CPTR9_normalized_dsp_object.RDS"</span>)</span>
<span id="cb2-4"><a href="#cb2-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb2-5"><a href="#cb2-5" aria-hidden="true" tabindex="-1"></a>normalized.object</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>NanoStringGeoMxSet (storageMode: lockedEnvironment)
assayData: 19704 features, 153 samples
element names: exprs, neg_norm, q_norm
protocolData
sampleNames: DSP-1001660013739-B-A02.dcc DSP-1001660013739-B-A03.dcc
... DSP-1001660022226-A-H12.dcc (153 total)
varLabels: FileVersion SoftwareVersion ... NegGeoMean (24 total)
varMetadata: labelDescription
phenoData
sampleNames: DSP-1001660013739-B-A02.dcc DSP-1001660013739-B-A03.dcc
... DSP-1001660022226-A-H12.dcc (153 total)
varLabels: slide_name nuclei ... neg_norm_negFactors (19 total)
varMetadata: labelDescription
featureData
featureNames: 0610009B22Rik Sanbr ... Trgc4 (19704 total)
fvarLabels: TargetName Module ... DetectionThreshold (9 total)
fvarMetadata: labelDescription
experimentData: use 'experimentData(object)'
Annotation: Mm_R_NGS_WTA_v1.0.pkc
signature: none
feature: Target
analyte: RNA</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb4"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb4-1"><a href="#cb4-1" aria-hidden="true" tabindex="-1"></a><span class="fu">dim</span>(normalized.object)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>Features Samples
19704 153 </code></pre>
</div>
</div>
<section id="differential-expression" class="level2">
<h2 class="anchored" data-anchor-id="differential-expression">Differential Expression</h2>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb6"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb6-1"><a href="#cb6-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Set up annotation colors for the heatmaps</span></span>
<span id="cb6-2"><a href="#cb6-2" aria-hidden="true" tabindex="-1"></a>anno.colors <span class="ot">=</span> <span class="fu">list</span>(</span>
<span id="cb6-3"><a href="#cb6-3" aria-hidden="true" tabindex="-1"></a> <span class="at">cell_type =</span> <span class="fu">c</span>(<span class="at">epithelial =</span> <span class="st">"slateblue"</span>, </span>
<span id="cb6-4"><a href="#cb6-4" aria-hidden="true" tabindex="-1"></a> <span class="at">myeloid =</span> <span class="st">"coral"</span>, </span>
<span id="cb6-5"><a href="#cb6-5" aria-hidden="true" tabindex="-1"></a> <span class="at">other =</span> <span class="st">"green"</span>), </span>
<span id="cb6-6"><a href="#cb6-6" aria-hidden="true" tabindex="-1"></a> <span class="at">region =</span> <span class="fu">c</span>(<span class="at">tumor =</span> <span class="st">"pink"</span>,</span>
<span id="cb6-7"><a href="#cb6-7" aria-hidden="true" tabindex="-1"></a> <span class="at">interstitial =</span> <span class="st">"seagreen1"</span>, </span>
<span id="cb6-8"><a href="#cb6-8" aria-hidden="true" tabindex="-1"></a> <span class="at">healthyepithelium =</span> <span class="st">"cadetblue1"</span>,</span>
<span id="cb6-9"><a href="#cb6-9" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster =</span> <span class="st">"darkorange"</span>), </span>
<span id="cb6-10"><a href="#cb6-10" aria-hidden="true" tabindex="-1"></a> <span class="at">segment =</span> <span class="fu">c</span>(<span class="at">PanCK =</span> <span class="st">"lightsalmon"</span>, </span>
<span id="cb6-11"><a href="#cb6-11" aria-hidden="true" tabindex="-1"></a> <span class="at">Myeloid =</span> <span class="st">"darkolivegreen1"</span>, </span>
<span id="cb6-12"><a href="#cb6-12" aria-hidden="true" tabindex="-1"></a> <span class="at">other_cells =</span> <span class="st">"lightblue"</span>)</span>
<span id="cb6-13"><a href="#cb6-13" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb6-14"><a href="#cb6-14" aria-hidden="true" tabindex="-1"></a> )</span>
<span id="cb6-15"><a href="#cb6-15" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb6-16"><a href="#cb6-16" aria-hidden="true" tabindex="-1"></a>DE.results.folder <span class="ot"><-</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>)</span>
<span id="cb6-17"><a href="#cb6-17" aria-hidden="true" tabindex="-1"></a>DE.results.files <span class="ot"><-</span> <span class="fu">list.files</span>(DE.results.folder)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
</div>
<section id="de-contrast-myeloid-vs.-panck" class="level3">
<h3 class="anchored" data-anchor-id="de-contrast-myeloid-vs.-panck">DE contrast: Myeloid vs. PanCK</h3>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb7"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb7-1"><a href="#cb7-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Setup annotation groups for the contrast</span></span>
<span id="cb7-2"><a href="#cb7-2" aria-hidden="true" tabindex="-1"></a>segment.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>, <span class="st">"PanCK"</span>)</span>
<span id="cb7-3"><a href="#cb7-3" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-4"><a href="#cb7-4" aria-hidden="true" tabindex="-1"></a><span class="co"># Define the name of the contrast</span></span>
<span id="cb7-5"><a href="#cb7-5" aria-hidden="true" tabindex="-1"></a>contrast.name <span class="ot"><-</span> <span class="fu">paste0</span>(segment.types[[<span class="dv">1</span>]], </span>
<span id="cb7-6"><a href="#cb7-6" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb7-7"><a href="#cb7-7" aria-hidden="true" tabindex="-1"></a> segment.types[[<span class="dv">2</span>]])</span>
<span id="cb7-8"><a href="#cb7-8" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-9"><a href="#cb7-9" aria-hidden="true" tabindex="-1"></a><span class="co"># Gather the column names to be used in defining contrasts</span></span>
<span id="cb7-10"><a href="#cb7-10" aria-hidden="true" tabindex="-1"></a>contrast.groups.list <span class="ot"><-</span> <span class="fu">list</span>(</span>
<span id="cb7-11"><a href="#cb7-11" aria-hidden="true" tabindex="-1"></a> <span class="st">"segment"</span> <span class="ot">=</span> segment.types)</span>
<span id="cb7-12"><a href="#cb7-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-13"><a href="#cb7-13" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-14"><a href="#cb7-14" aria-hidden="true" tabindex="-1"></a><span class="co"># Filter data for the chosen annotations</span></span>
<span id="cb7-15"><a href="#cb7-15" aria-hidden="true" tabindex="-1"></a>object <span class="ot"><-</span> normalized.object</span>
<span id="cb7-16"><a href="#cb7-16" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-17"><a href="#cb7-17" aria-hidden="true" tabindex="-1"></a><span class="co"># Generate the counts, annotation, and subset object for lmm</span></span>
<span id="cb7-18"><a href="#cb7-18" aria-hidden="true" tabindex="-1"></a>lmm.input <span class="ot"><-</span> <span class="fu">subset_for_lmm</span>(<span class="at">object =</span> object, </span>
<span id="cb7-19"><a href="#cb7-19" aria-hidden="true" tabindex="-1"></a> <span class="at">subset.list =</span> contrast.groups.list)</span>
<span id="cb7-20"><a href="#cb7-20" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-21"><a href="#cb7-21" aria-hidden="true" tabindex="-1"></a><span class="co"># Create summary table of group counts</span></span>
<span id="cb7-22"><a href="#cb7-22" aria-hidden="true" tabindex="-1"></a>summary.table.df <span class="ot"><-</span> <span class="fu">pData</span>(lmm.input<span class="sc">$</span>subset.object) <span class="sc">%>%</span> </span>
<span id="cb7-23"><a href="#cb7-23" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">c</span>(<span class="fu">names</span>(contrast.groups.list)))</span>
<span id="cb7-24"><a href="#cb7-24" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-25"><a href="#cb7-25" aria-hidden="true" tabindex="-1"></a>summary.table <span class="ot"><-</span> <span class="fu">table</span>(summary.table.df)</span>
<span id="cb7-26"><a href="#cb7-26" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb7-27"><a href="#cb7-27" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"Sample Numbers per Annotation Group"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "Sample Numbers per Annotation Group"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb9"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb9-1"><a href="#cb9-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"-----------------------------------"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "-----------------------------------"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb11"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb11-1"><a href="#cb11-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(summary.table)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>segment
Myeloid PanCK
57 36 </code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb13"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb13-1"><a href="#cb13-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Check if the DE results have already been generated</span></span>
<span id="cb13-2"><a href="#cb13-2" aria-hidden="true" tabindex="-1"></a>DE.result.file <span class="ot"><-</span> <span class="fu">grepl</span>(contrast.name, DE.results.files)</span>
<span id="cb13-3"><a href="#cb13-3" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(<span class="fu">sum</span>(DE.result.file) <span class="sc">==</span> <span class="dv">1</span>) {</span>
<span id="cb13-4"><a href="#cb13-4" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb13-5"><a href="#cb13-5" aria-hidden="true" tabindex="-1"></a> <span class="co"># Load the previously generated DE results</span></span>
<span id="cb13-6"><a href="#cb13-6" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> <span class="fu">as.data.frame</span>(<span class="fu">read.csv</span>(<span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>)))</span>
<span id="cb13-7"><a href="#cb13-7" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb13-8"><a href="#cb13-8" aria-hidden="true" tabindex="-1"></a>} <span class="cf">else</span> {</span>
<span id="cb13-9"><a href="#cb13-9" aria-hidden="true" tabindex="-1"></a> <span class="co"># Use the function from the DSPWorkflow package</span></span>
<span id="cb13-10"><a href="#cb13-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb13-11"><a href="#cb13-11" aria-hidden="true" tabindex="-1"></a> <span class="co"># Within slide analysis</span></span>
<span id="cb13-12"><a href="#cb13-12" aria-hidden="true" tabindex="-1"></a> <span class="co"># Listed contrasts are condition, reference </span></span>
<span id="cb13-13"><a href="#cb13-13" aria-hidden="true" tabindex="-1"></a> results.list <span class="ot"><-</span> <span class="fu">diffExpr</span>(<span class="at">object =</span> lmm.input<span class="sc">$</span>subset.object, </span>
<span id="cb13-14"><a href="#cb13-14" aria-hidden="true" tabindex="-1"></a> <span class="at">analysis.type =</span> <span class="st">"Within Groups"</span>, </span>
<span id="cb13-15"><a href="#cb13-15" aria-hidden="true" tabindex="-1"></a> <span class="at">region.col =</span> <span class="st">"segment"</span>, </span>
<span id="cb13-16"><a href="#cb13-16" aria-hidden="true" tabindex="-1"></a> <span class="at">regions =</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>, <span class="st">"PanCK"</span>), </span>
<span id="cb13-17"><a href="#cb13-17" aria-hidden="true" tabindex="-1"></a> <span class="at">group.col =</span> <span class="st">"class"</span>, </span>
<span id="cb13-18"><a href="#cb13-18" aria-hidden="true" tabindex="-1"></a> <span class="at">groups =</span> <span class="fu">c</span>(<span class="st">"cancer"</span>), </span>
<span id="cb13-19"><a href="#cb13-19" aria-hidden="true" tabindex="-1"></a> <span class="at">n.cores =</span> parallel<span class="sc">::</span><span class="fu">detectCores</span>())</span>
<span id="cb13-20"><a href="#cb13-20" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb13-21"><a href="#cb13-21" aria-hidden="true" tabindex="-1"></a> <span class="co"># Create the results df</span></span>
<span id="cb13-22"><a href="#cb13-22" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> results.list<span class="sc">$</span>results</span>
<span id="cb13-23"><a href="#cb13-23" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb13-24"><a href="#cb13-24" aria-hidden="true" tabindex="-1"></a> write.results <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb13-25"><a href="#cb13-25" aria-hidden="true" tabindex="-1"></a> <span class="cf">if</span>(write.results <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb13-26"><a href="#cb13-26" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>))</span>
<span id="cb13-27"><a href="#cb13-27" aria-hidden="true" tabindex="-1"></a> }</span>
<span id="cb13-28"><a href="#cb13-28" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb13-29"><a href="#cb13-29" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb13-30"><a href="#cb13-30" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>Running Within Group Analysis between Regions
Number of regions in group cancer: 2
[1] "Running DE with linear model (no mixed-effect)"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb15"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb15-1"><a href="#cb15-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Adjust column names</span></span>
<span id="cb15-2"><a href="#cb15-2" aria-hidden="true" tabindex="-1"></a>logfc.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"logFC"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb15-3"><a href="#cb15-3" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>logfc <span class="ot"><-</span> results.df[[logfc.column]]</span>
<span id="cb15-4"><a href="#cb15-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-5"><a href="#cb15-5" aria-hidden="true" tabindex="-1"></a>pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"_pval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb15-6"><a href="#cb15-6" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>pval <span class="ot"><-</span> results.df[[pval.column]]</span>
<span id="cb15-7"><a href="#cb15-7" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-8"><a href="#cb15-8" aria-hidden="true" tabindex="-1"></a>adj.pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"adjpval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb15-9"><a href="#cb15-9" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>padj <span class="ot"><-</span> results.df[[adj.pval.column]]</span>
<span id="cb15-10"><a href="#cb15-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-11"><a href="#cb15-11" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>gene <span class="ot"><-</span> results.df<span class="sc">$</span>Gene</span>
<span id="cb15-12"><a href="#cb15-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-13"><a href="#cb15-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Keep only the necessary columns</span></span>
<span id="cb15-14"><a href="#cb15-14" aria-hidden="true" tabindex="-1"></a>results.df <span class="ot"><-</span> results.df <span class="sc">%>%</span> <span class="fu">select</span>(<span class="fu">c</span>(<span class="st">"gene"</span>, </span>
<span id="cb15-15"><a href="#cb15-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"logfc"</span>, </span>
<span id="cb15-16"><a href="#cb15-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"pval"</span>, </span>
<span id="cb15-17"><a href="#cb15-17" aria-hidden="true" tabindex="-1"></a> <span class="st">"padj"</span>))</span>
<span id="cb15-18"><a href="#cb15-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-19"><a href="#cb15-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-20"><a href="#cb15-20" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the results</span></span>
<span id="cb15-21"><a href="#cb15-21" aria-hidden="true" tabindex="-1"></a>export.deg.list <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb15-22"><a href="#cb15-22" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.deg.list <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb15-23"><a href="#cb15-23" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, </span>
<span id="cb15-24"><a href="#cb15-24" aria-hidden="true" tabindex="-1"></a> <span class="at">file =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/DEG_lists/"</span>, contrast.name, <span class="st">"_deg_list.csv"</span>), </span>
<span id="cb15-25"><a href="#cb15-25" aria-hidden="true" tabindex="-1"></a> <span class="at">row.names =</span> <span class="cn">FALSE</span>) </span>
<span id="cb15-26"><a href="#cb15-26" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb15-27"><a href="#cb15-27" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-28"><a href="#cb15-28" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the volcano plot</span></span>
<span id="cb15-29"><a href="#cb15-29" aria-hidden="true" tabindex="-1"></a>volcano.output <span class="ot"><-</span> <span class="fu">make_volcano</span>(<span class="at">lmm.results =</span> results.df, </span>
<span id="cb15-30"><a href="#cb15-30" aria-hidden="true" tabindex="-1"></a> <span class="at">title =</span> contrast.name, </span>
<span id="cb15-31"><a href="#cb15-31" aria-hidden="true" tabindex="-1"></a> <span class="at">legend.title =</span> <span class="st">"Expression in Myeloid"</span>, </span>
<span id="cb15-32"><a href="#cb15-32" aria-hidden="true" tabindex="-1"></a> <span class="at">x.axis.title =</span> <span class="st">"Myeloid (cond) versus PanCK (ref)"</span>, </span>
<span id="cb15-33"><a href="#cb15-33" aria-hidden="true" tabindex="-1"></a> <span class="at">fc.limit =</span> <span class="dv">1</span>, </span>
<span id="cb15-34"><a href="#cb15-34" aria-hidden="true" tabindex="-1"></a> <span class="at">pos.label.limit =</span> <span class="dv">3</span>, </span>
<span id="cb15-35"><a href="#cb15-35" aria-hidden="true" tabindex="-1"></a> <span class="at">neg.label.limit =</span> <span class="sc">-</span><span class="dv">1</span>)</span>
<span id="cb15-36"><a href="#cb15-36" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-37"><a href="#cb15-37" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the volcano</span></span>
<span id="cb15-38"><a href="#cb15-38" aria-hidden="true" tabindex="-1"></a>export.volcano <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb15-39"><a href="#cb15-39" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.volcano <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb15-40"><a href="#cb15-40" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(<span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/volcano/"</span>, contrast.name, <span class="st">"_volcano_plot.png"</span>), </span>
<span id="cb15-41"><a href="#cb15-41" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb15-42"><a href="#cb15-42" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb15-43"><a href="#cb15-43" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb15-44"><a href="#cb15-44" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb15-45"><a href="#cb15-45" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(volcano.output<span class="sc">$</span>volcano.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast-%20Myeloid%20vs.%20PanCK-1.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb16"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb16-1"><a href="#cb16-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the heatmap</span></span>
<span id="cb16-2"><a href="#cb16-2" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb16-3"><a href="#cb16-3" aria-hidden="true" tabindex="-1"></a><span class="co"># Define new annotations if needed</span></span>
<span id="cb16-4"><a href="#cb16-4" aria-hidden="true" tabindex="-1"></a>anno.colors <span class="ot">=</span> <span class="fu">list</span>(<span class="at">region =</span> <span class="fu">c</span>(<span class="at">tumor =</span> <span class="st">"pink"</span>,</span>
<span id="cb16-5"><a href="#cb16-5" aria-hidden="true" tabindex="-1"></a> <span class="at">interstitial =</span> <span class="st">"seagreen1"</span>, </span>
<span id="cb16-6"><a href="#cb16-6" aria-hidden="true" tabindex="-1"></a> <span class="at">healthyepithelium =</span> <span class="st">"cadetblue1"</span>,</span>
<span id="cb16-7"><a href="#cb16-7" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster =</span> <span class="st">"darkorange"</span>), </span>
<span id="cb16-8"><a href="#cb16-8" aria-hidden="true" tabindex="-1"></a> <span class="at">segment =</span> <span class="fu">c</span>(<span class="at">PanCK =</span> <span class="st">"lightsalmon"</span>, </span>
<span id="cb16-9"><a href="#cb16-9" aria-hidden="true" tabindex="-1"></a> <span class="at">Myeloid =</span> <span class="st">"darkolivegreen1"</span>, </span>
<span id="cb16-10"><a href="#cb16-10" aria-hidden="true" tabindex="-1"></a> <span class="at">other_cells =</span> <span class="st">"lightblue"</span>))</span>
<span id="cb16-11"><a href="#cb16-11" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb16-12"><a href="#cb16-12" aria-hidden="true" tabindex="-1"></a><span class="co"># Subset the annotation just for the heatmap annotations of interest</span></span>
<span id="cb16-13"><a href="#cb16-13" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> lmm.input<span class="sc">$</span>annotation <span class="sc">%>%</span> </span>
<span id="cb16-14"><a href="#cb16-14" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">names</span>(anno.colors))</span>
<span id="cb16-15"><a href="#cb16-15" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb16-16"><a href="#cb16-16" aria-hidden="true" tabindex="-1"></a><span class="co"># Order the annotation</span></span>
<span id="cb16-17"><a href="#cb16-17" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>region), ]</span>
<span id="cb16-18"><a href="#cb16-18" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>segment), ]</span>
<span id="cb16-19"><a href="#cb16-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb16-20"><a href="#cb16-20" aria-hidden="true" tabindex="-1"></a>heatmap.plot <span class="ot"><-</span> <span class="fu">make_heatmap</span>(</span>
<span id="cb16-21"><a href="#cb16-21" aria-hidden="true" tabindex="-1"></a> <span class="at">normalized.log.counts.df =</span> lmm.input<span class="sc">$</span>log.counts, </span>
<span id="cb16-22"><a href="#cb16-22" aria-hidden="true" tabindex="-1"></a> <span class="at">de.results =</span> results.df, </span>
<span id="cb16-23"><a href="#cb16-23" aria-hidden="true" tabindex="-1"></a> <span class="at">top.degs =</span> <span class="cn">TRUE</span>, </span>
<span id="cb16-24"><a href="#cb16-24" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.column =</span> annotation.heatmap,</span>
<span id="cb16-25"><a href="#cb16-25" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.row =</span> <span class="cn">NULL</span>, </span>
<span id="cb16-26"><a href="#cb16-26" aria-hidden="true" tabindex="-1"></a> <span class="at">anno.colors =</span> anno.colors, </span>
<span id="cb16-27"><a href="#cb16-27" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.rows =</span> <span class="cn">TRUE</span>, </span>
<span id="cb16-28"><a href="#cb16-28" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.columns =</span> <span class="cn">TRUE</span>, </span>
<span id="cb16-29"><a href="#cb16-29" aria-hidden="true" tabindex="-1"></a> <span class="at">main.title =</span> contrast.name, </span>
<span id="cb16-30"><a href="#cb16-30" aria-hidden="true" tabindex="-1"></a> <span class="at">row.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb16-31"><a href="#cb16-31" aria-hidden="true" tabindex="-1"></a> <span class="at">column.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb16-32"><a href="#cb16-32" aria-hidden="true" tabindex="-1"></a> <span class="at">show.colnames =</span> <span class="cn">FALSE</span>)</span>
<span id="cb16-33"><a href="#cb16-33" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb16-34"><a href="#cb16-34" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(heatmap.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast-%20Myeloid%20vs.%20PanCK-2.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb17"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb17-1"><a href="#cb17-1" aria-hidden="true" tabindex="-1"></a>export.heatmap <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb17-2"><a href="#cb17-2" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.heatmap <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb17-3"><a href="#cb17-3" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb17-4"><a href="#cb17-4" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(heatmap.plot, </span>
<span id="cb17-5"><a href="#cb17-5" aria-hidden="true" tabindex="-1"></a> <span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/heatmap/"</span>, contrast.name, <span class="st">"_heatmap_plot.png"</span>), </span>
<span id="cb17-6"><a href="#cb17-6" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb17-7"><a href="#cb17-7" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb17-8"><a href="#cb17-8" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb17-9"><a href="#cb17-9" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
</div>
</section>
<section id="de-contrast-panck-tumor-vs-panck-healthy-epithelium" class="level3">
<h3 class="anchored" data-anchor-id="de-contrast-panck-tumor-vs-panck-healthy-epithelium">DE contrast: PanCK tumor vs PanCK healthy epithelium</h3>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb18"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb18-1"><a href="#cb18-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Setup annotation groups for the contrast</span></span>
<span id="cb18-2"><a href="#cb18-2" aria-hidden="true" tabindex="-1"></a>segment.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"PanCK"</span>)</span>
<span id="cb18-3"><a href="#cb18-3" aria-hidden="true" tabindex="-1"></a>region.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"tumor"</span>, <span class="st">"healthyepithelium"</span>)</span>
<span id="cb18-4"><a href="#cb18-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-5"><a href="#cb18-5" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-6"><a href="#cb18-6" aria-hidden="true" tabindex="-1"></a><span class="co"># Define the name of the contrast</span></span>
<span id="cb18-7"><a href="#cb18-7" aria-hidden="true" tabindex="-1"></a>contrast.name <span class="ot"><-</span> <span class="fu">paste0</span>(segment.types[[<span class="dv">1</span>]], </span>
<span id="cb18-8"><a href="#cb18-8" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb18-9"><a href="#cb18-9" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">1</span>]], </span>
<span id="cb18-10"><a href="#cb18-10" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb18-11"><a href="#cb18-11" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">2</span>]])</span>
<span id="cb18-12"><a href="#cb18-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-13"><a href="#cb18-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Gather the column names to be used in defining contrasts</span></span>
<span id="cb18-14"><a href="#cb18-14" aria-hidden="true" tabindex="-1"></a>contrast.groups.list <span class="ot"><-</span> <span class="fu">list</span>(</span>
<span id="cb18-15"><a href="#cb18-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"segment"</span> <span class="ot">=</span> segment.types, </span>
<span id="cb18-16"><a href="#cb18-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"region"</span> <span class="ot">=</span> region.types)</span>
<span id="cb18-17"><a href="#cb18-17" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-18"><a href="#cb18-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-19"><a href="#cb18-19" aria-hidden="true" tabindex="-1"></a><span class="co"># Filter data for the chosen annotations</span></span>
<span id="cb18-20"><a href="#cb18-20" aria-hidden="true" tabindex="-1"></a>object <span class="ot"><-</span> normalized.object</span>
<span id="cb18-21"><a href="#cb18-21" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-22"><a href="#cb18-22" aria-hidden="true" tabindex="-1"></a><span class="co"># Generate the counts, annotation, and subset object for lmm</span></span>
<span id="cb18-23"><a href="#cb18-23" aria-hidden="true" tabindex="-1"></a>lmm.input <span class="ot"><-</span> <span class="fu">subset_for_lmm</span>(<span class="at">object =</span> object, </span>
<span id="cb18-24"><a href="#cb18-24" aria-hidden="true" tabindex="-1"></a> <span class="at">subset.list =</span> contrast.groups.list)</span>
<span id="cb18-25"><a href="#cb18-25" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-26"><a href="#cb18-26" aria-hidden="true" tabindex="-1"></a><span class="co"># Create summary table of group counts</span></span>
<span id="cb18-27"><a href="#cb18-27" aria-hidden="true" tabindex="-1"></a>summary.table.df <span class="ot"><-</span> <span class="fu">pData</span>(lmm.input<span class="sc">$</span>subset.object) <span class="sc">%>%</span> </span>
<span id="cb18-28"><a href="#cb18-28" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">c</span>(<span class="fu">names</span>(contrast.groups.list)))</span>
<span id="cb18-29"><a href="#cb18-29" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-30"><a href="#cb18-30" aria-hidden="true" tabindex="-1"></a>summary.table <span class="ot"><-</span> <span class="fu">table</span>(summary.table.df)</span>
<span id="cb18-31"><a href="#cb18-31" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb18-32"><a href="#cb18-32" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"Sample Numbers per Annotation Group"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "Sample Numbers per Annotation Group"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb20"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb20-1"><a href="#cb20-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"-----------------------------------"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "-----------------------------------"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb22"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb22-1"><a href="#cb22-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(summary.table)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code> region
segment healthyepithelium tumor
PanCK 15 19</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb24"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb24-1"><a href="#cb24-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Check if the DE results have already been generated</span></span>
<span id="cb24-2"><a href="#cb24-2" aria-hidden="true" tabindex="-1"></a>DE.result.file <span class="ot"><-</span> <span class="fu">grepl</span>(contrast.name, DE.results.files)</span>
<span id="cb24-3"><a href="#cb24-3" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(<span class="fu">sum</span>(DE.result.file) <span class="sc">==</span> <span class="dv">1</span>) {</span>
<span id="cb24-4"><a href="#cb24-4" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb24-5"><a href="#cb24-5" aria-hidden="true" tabindex="-1"></a> <span class="co"># Load the previously generated DE results</span></span>
<span id="cb24-6"><a href="#cb24-6" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> <span class="fu">as.data.frame</span>(<span class="fu">read.csv</span>(<span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>)))</span>
<span id="cb24-7"><a href="#cb24-7" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb24-8"><a href="#cb24-8" aria-hidden="true" tabindex="-1"></a>} <span class="cf">else</span> {</span>
<span id="cb24-9"><a href="#cb24-9" aria-hidden="true" tabindex="-1"></a> <span class="co"># Use the function from the DSPWorkflow package</span></span>
<span id="cb24-10"><a href="#cb24-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb24-11"><a href="#cb24-11" aria-hidden="true" tabindex="-1"></a> <span class="co"># Within slide analysis</span></span>
<span id="cb24-12"><a href="#cb24-12" aria-hidden="true" tabindex="-1"></a> <span class="co"># Listed contrasts are condition, reference </span></span>
<span id="cb24-13"><a href="#cb24-13" aria-hidden="true" tabindex="-1"></a> results.list <span class="ot"><-</span> <span class="fu">diffExpr</span>(<span class="at">object =</span> lmm.input<span class="sc">$</span>subset.object, </span>
<span id="cb24-14"><a href="#cb24-14" aria-hidden="true" tabindex="-1"></a> <span class="at">analysis.type =</span> <span class="st">"Within Groups"</span>, </span>
<span id="cb24-15"><a href="#cb24-15" aria-hidden="true" tabindex="-1"></a> <span class="at">region.col =</span> <span class="st">"region"</span>, </span>
<span id="cb24-16"><a href="#cb24-16" aria-hidden="true" tabindex="-1"></a> <span class="at">regions =</span> <span class="fu">c</span>(<span class="st">"tumor"</span>, <span class="st">"healthyepithelium"</span>), </span>
<span id="cb24-17"><a href="#cb24-17" aria-hidden="true" tabindex="-1"></a> <span class="at">group.col =</span> <span class="st">"segment"</span>, </span>
<span id="cb24-18"><a href="#cb24-18" aria-hidden="true" tabindex="-1"></a> <span class="at">groups =</span> <span class="fu">c</span>(<span class="st">"PanCK"</span>), </span>
<span id="cb24-19"><a href="#cb24-19" aria-hidden="true" tabindex="-1"></a> <span class="at">n.cores =</span> parallel<span class="sc">::</span><span class="fu">detectCores</span>())</span>
<span id="cb24-20"><a href="#cb24-20" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb24-21"><a href="#cb24-21" aria-hidden="true" tabindex="-1"></a> <span class="co"># Create the results df</span></span>
<span id="cb24-22"><a href="#cb24-22" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> results.list<span class="sc">$</span>results</span>
<span id="cb24-23"><a href="#cb24-23" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb24-24"><a href="#cb24-24" aria-hidden="true" tabindex="-1"></a> write.results <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb24-25"><a href="#cb24-25" aria-hidden="true" tabindex="-1"></a> <span class="cf">if</span>(write.results <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb24-26"><a href="#cb24-26" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>))</span>
<span id="cb24-27"><a href="#cb24-27" aria-hidden="true" tabindex="-1"></a> }</span>
<span id="cb24-28"><a href="#cb24-28" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb24-29"><a href="#cb24-29" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>Running Within Group Analysis between Regions
Number of regions in group PanCK: 2
[1] "Running DE with linear model (no mixed-effect)"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb26"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb26-1"><a href="#cb26-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Adjust column names</span></span>
<span id="cb26-2"><a href="#cb26-2" aria-hidden="true" tabindex="-1"></a>logfc.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"logFC"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb26-3"><a href="#cb26-3" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>logfc <span class="ot"><-</span> results.df[[logfc.column]]</span>
<span id="cb26-4"><a href="#cb26-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-5"><a href="#cb26-5" aria-hidden="true" tabindex="-1"></a>pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"_pval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb26-6"><a href="#cb26-6" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>pval <span class="ot"><-</span> results.df[[pval.column]]</span>
<span id="cb26-7"><a href="#cb26-7" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-8"><a href="#cb26-8" aria-hidden="true" tabindex="-1"></a>adj.pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"adjpval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb26-9"><a href="#cb26-9" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>padj <span class="ot"><-</span> results.df[[adj.pval.column]]</span>
<span id="cb26-10"><a href="#cb26-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-11"><a href="#cb26-11" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>gene <span class="ot"><-</span> results.df<span class="sc">$</span>Gene</span>
<span id="cb26-12"><a href="#cb26-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-13"><a href="#cb26-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Keep only the necessary columns</span></span>
<span id="cb26-14"><a href="#cb26-14" aria-hidden="true" tabindex="-1"></a>results.df <span class="ot"><-</span> results.df <span class="sc">%>%</span> <span class="fu">select</span>(<span class="fu">c</span>(<span class="st">"gene"</span>, </span>
<span id="cb26-15"><a href="#cb26-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"logfc"</span>, </span>
<span id="cb26-16"><a href="#cb26-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"pval"</span>, </span>
<span id="cb26-17"><a href="#cb26-17" aria-hidden="true" tabindex="-1"></a> <span class="st">"padj"</span>))</span>
<span id="cb26-18"><a href="#cb26-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-19"><a href="#cb26-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-20"><a href="#cb26-20" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the results</span></span>
<span id="cb26-21"><a href="#cb26-21" aria-hidden="true" tabindex="-1"></a>export.deg.list <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb26-22"><a href="#cb26-22" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.deg.list <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb26-23"><a href="#cb26-23" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, </span>
<span id="cb26-24"><a href="#cb26-24" aria-hidden="true" tabindex="-1"></a> <span class="at">file =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/DEG_lists/"</span>, contrast.name, <span class="st">"_deg_list.csv"</span>), </span>
<span id="cb26-25"><a href="#cb26-25" aria-hidden="true" tabindex="-1"></a> <span class="at">row.names =</span> <span class="cn">FALSE</span>) </span>
<span id="cb26-26"><a href="#cb26-26" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb26-27"><a href="#cb26-27" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-28"><a href="#cb26-28" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the volcano plot</span></span>
<span id="cb26-29"><a href="#cb26-29" aria-hidden="true" tabindex="-1"></a>volcano.output <span class="ot"><-</span> <span class="fu">make_volcano</span>(<span class="at">lmm.results =</span> results.df, </span>
<span id="cb26-30"><a href="#cb26-30" aria-hidden="true" tabindex="-1"></a> <span class="at">title =</span> contrast.name, </span>
<span id="cb26-31"><a href="#cb26-31" aria-hidden="true" tabindex="-1"></a> <span class="at">legend.title =</span> <span class="st">"Expression in Tumor"</span>, </span>
<span id="cb26-32"><a href="#cb26-32" aria-hidden="true" tabindex="-1"></a> <span class="at">x.axis.title =</span> <span class="st">"Tumor (cond) versus Healthy Epi (ref)"</span>, </span>
<span id="cb26-33"><a href="#cb26-33" aria-hidden="true" tabindex="-1"></a> <span class="at">fc.limit =</span> <span class="dv">1</span>, </span>
<span id="cb26-34"><a href="#cb26-34" aria-hidden="true" tabindex="-1"></a> <span class="at">pos.label.limit =</span> <span class="dv">1</span>, </span>
<span id="cb26-35"><a href="#cb26-35" aria-hidden="true" tabindex="-1"></a> <span class="at">neg.label.limit =</span> <span class="sc">-</span><span class="dv">1</span>)</span>
<span id="cb26-36"><a href="#cb26-36" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-37"><a href="#cb26-37" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the volcano</span></span>
<span id="cb26-38"><a href="#cb26-38" aria-hidden="true" tabindex="-1"></a>export.volcano <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb26-39"><a href="#cb26-39" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.volcano <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb26-40"><a href="#cb26-40" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(<span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/volcano/"</span>, contrast.name, <span class="st">"_volcano_plot.png"</span>), </span>
<span id="cb26-41"><a href="#cb26-41" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb26-42"><a href="#cb26-42" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb26-43"><a href="#cb26-43" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb26-44"><a href="#cb26-44" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb26-45"><a href="#cb26-45" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(volcano.output<span class="sc">$</span>volcano.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast%20PanCK%20tumor%20vs%20PanCK%20healthy%20epithelium-1.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb27"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb27-1"><a href="#cb27-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the heatmap</span></span>
<span id="cb27-2"><a href="#cb27-2" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb27-3"><a href="#cb27-3" aria-hidden="true" tabindex="-1"></a><span class="co"># Define new annotations if needed</span></span>
<span id="cb27-4"><a href="#cb27-4" aria-hidden="true" tabindex="-1"></a>anno.colors <span class="ot">=</span> <span class="fu">list</span>(<span class="at">region =</span> <span class="fu">c</span>(<span class="at">tumor =</span> <span class="st">"pink"</span>,</span>
<span id="cb27-5"><a href="#cb27-5" aria-hidden="true" tabindex="-1"></a> <span class="at">interstitial =</span> <span class="st">"seagreen1"</span>, </span>
<span id="cb27-6"><a href="#cb27-6" aria-hidden="true" tabindex="-1"></a> <span class="at">healthyepithelium =</span> <span class="st">"cadetblue1"</span>,</span>
<span id="cb27-7"><a href="#cb27-7" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster =</span> <span class="st">"darkorange"</span>), </span>
<span id="cb27-8"><a href="#cb27-8" aria-hidden="true" tabindex="-1"></a> <span class="at">segment =</span> <span class="fu">c</span>(<span class="at">PanCK =</span> <span class="st">"lightsalmon"</span>, </span>
<span id="cb27-9"><a href="#cb27-9" aria-hidden="true" tabindex="-1"></a> <span class="at">Myeloid =</span> <span class="st">"darkolivegreen1"</span>, </span>
<span id="cb27-10"><a href="#cb27-10" aria-hidden="true" tabindex="-1"></a> <span class="at">other_cells =</span> <span class="st">"lightblue"</span>))</span>
<span id="cb27-11"><a href="#cb27-11" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb27-12"><a href="#cb27-12" aria-hidden="true" tabindex="-1"></a><span class="co"># Subset the annotation just for the heatmap annotations of interest</span></span>
<span id="cb27-13"><a href="#cb27-13" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> lmm.input<span class="sc">$</span>annotation <span class="sc">%>%</span> </span>
<span id="cb27-14"><a href="#cb27-14" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">names</span>(anno.colors))</span>
<span id="cb27-15"><a href="#cb27-15" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb27-16"><a href="#cb27-16" aria-hidden="true" tabindex="-1"></a><span class="co"># Order the annotation</span></span>
<span id="cb27-17"><a href="#cb27-17" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>region), ]</span>
<span id="cb27-18"><a href="#cb27-18" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>segment), ]</span>
<span id="cb27-19"><a href="#cb27-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb27-20"><a href="#cb27-20" aria-hidden="true" tabindex="-1"></a>heatmap.plot <span class="ot"><-</span> <span class="fu">make_heatmap</span>(</span>
<span id="cb27-21"><a href="#cb27-21" aria-hidden="true" tabindex="-1"></a> <span class="at">normalized.log.counts.df =</span> lmm.input<span class="sc">$</span>log.counts, </span>
<span id="cb27-22"><a href="#cb27-22" aria-hidden="true" tabindex="-1"></a> <span class="at">de.results =</span> results.df, </span>
<span id="cb27-23"><a href="#cb27-23" aria-hidden="true" tabindex="-1"></a> <span class="at">top.degs =</span> <span class="cn">TRUE</span>, </span>
<span id="cb27-24"><a href="#cb27-24" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.column =</span> annotation.heatmap,</span>
<span id="cb27-25"><a href="#cb27-25" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.row =</span> <span class="cn">NULL</span>, </span>
<span id="cb27-26"><a href="#cb27-26" aria-hidden="true" tabindex="-1"></a> <span class="at">anno.colors =</span> anno.colors, </span>
<span id="cb27-27"><a href="#cb27-27" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.rows =</span> <span class="cn">TRUE</span>, </span>
<span id="cb27-28"><a href="#cb27-28" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.columns =</span> <span class="cn">TRUE</span>, </span>
<span id="cb27-29"><a href="#cb27-29" aria-hidden="true" tabindex="-1"></a> <span class="at">main.title =</span> contrast.name, </span>
<span id="cb27-30"><a href="#cb27-30" aria-hidden="true" tabindex="-1"></a> <span class="at">row.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb27-31"><a href="#cb27-31" aria-hidden="true" tabindex="-1"></a> <span class="at">column.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb27-32"><a href="#cb27-32" aria-hidden="true" tabindex="-1"></a> <span class="at">show.colnames =</span> <span class="cn">FALSE</span>)</span>
<span id="cb27-33"><a href="#cb27-33" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb27-34"><a href="#cb27-34" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(heatmap.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast%20PanCK%20tumor%20vs%20PanCK%20healthy%20epithelium-2.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb28"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb28-1"><a href="#cb28-1" aria-hidden="true" tabindex="-1"></a>export.heatmap <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb28-2"><a href="#cb28-2" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.heatmap <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb28-3"><a href="#cb28-3" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb28-4"><a href="#cb28-4" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(heatmap.plot, </span>
<span id="cb28-5"><a href="#cb28-5" aria-hidden="true" tabindex="-1"></a> <span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/heatmap/"</span>, contrast.name, <span class="st">"_heatmap_plot.png"</span>), </span>
<span id="cb28-6"><a href="#cb28-6" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb28-7"><a href="#cb28-7" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb28-8"><a href="#cb28-8" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb28-9"><a href="#cb28-9" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
</div>
</section>
<section id="de-contrast-myeloid-tumor-vs-myeloid-interstitial" class="level3">
<h3 class="anchored" data-anchor-id="de-contrast-myeloid-tumor-vs-myeloid-interstitial">DE contrast: Myeloid Tumor vs Myeloid Interstitial</h3>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb29"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb29-1"><a href="#cb29-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Setup annotation groups for the contrast</span></span>
<span id="cb29-2"><a href="#cb29-2" aria-hidden="true" tabindex="-1"></a>segment.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>)</span>
<span id="cb29-3"><a href="#cb29-3" aria-hidden="true" tabindex="-1"></a>region.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"tumor"</span>, <span class="st">"interstitial"</span>)</span>
<span id="cb29-4"><a href="#cb29-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-5"><a href="#cb29-5" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-6"><a href="#cb29-6" aria-hidden="true" tabindex="-1"></a><span class="co"># Define the name of the contrast</span></span>
<span id="cb29-7"><a href="#cb29-7" aria-hidden="true" tabindex="-1"></a>contrast.name <span class="ot"><-</span> <span class="fu">paste0</span>(segment.types[[<span class="dv">1</span>]], </span>
<span id="cb29-8"><a href="#cb29-8" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb29-9"><a href="#cb29-9" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">1</span>]], </span>
<span id="cb29-10"><a href="#cb29-10" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb29-11"><a href="#cb29-11" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">2</span>]])</span>
<span id="cb29-12"><a href="#cb29-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-13"><a href="#cb29-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Gather the column names to be used in defining contrasts</span></span>
<span id="cb29-14"><a href="#cb29-14" aria-hidden="true" tabindex="-1"></a>contrast.groups.list <span class="ot"><-</span> <span class="fu">list</span>(</span>
<span id="cb29-15"><a href="#cb29-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"segment"</span> <span class="ot">=</span> segment.types, </span>
<span id="cb29-16"><a href="#cb29-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"region"</span> <span class="ot">=</span> region.types)</span>
<span id="cb29-17"><a href="#cb29-17" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-18"><a href="#cb29-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-19"><a href="#cb29-19" aria-hidden="true" tabindex="-1"></a><span class="co"># Filter data for the chosen annotations</span></span>
<span id="cb29-20"><a href="#cb29-20" aria-hidden="true" tabindex="-1"></a>object <span class="ot"><-</span> normalized.object</span>
<span id="cb29-21"><a href="#cb29-21" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-22"><a href="#cb29-22" aria-hidden="true" tabindex="-1"></a><span class="co"># Generate the counts, annotation, and subset object for lmm</span></span>
<span id="cb29-23"><a href="#cb29-23" aria-hidden="true" tabindex="-1"></a>lmm.input <span class="ot"><-</span> <span class="fu">subset_for_lmm</span>(<span class="at">object =</span> object, </span>
<span id="cb29-24"><a href="#cb29-24" aria-hidden="true" tabindex="-1"></a> <span class="at">subset.list =</span> contrast.groups.list)</span>
<span id="cb29-25"><a href="#cb29-25" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-26"><a href="#cb29-26" aria-hidden="true" tabindex="-1"></a><span class="co"># Create summary table of group counts</span></span>
<span id="cb29-27"><a href="#cb29-27" aria-hidden="true" tabindex="-1"></a>summary.table.df <span class="ot"><-</span> <span class="fu">pData</span>(lmm.input<span class="sc">$</span>subset.object) <span class="sc">%>%</span> </span>
<span id="cb29-28"><a href="#cb29-28" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">c</span>(<span class="fu">names</span>(contrast.groups.list)))</span>
<span id="cb29-29"><a href="#cb29-29" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-30"><a href="#cb29-30" aria-hidden="true" tabindex="-1"></a>summary.table <span class="ot"><-</span> <span class="fu">table</span>(summary.table.df)</span>
<span id="cb29-31"><a href="#cb29-31" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb29-32"><a href="#cb29-32" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"Sample Numbers per Annotation Group"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "Sample Numbers per Annotation Group"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb31"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb31-1"><a href="#cb31-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"-----------------------------------"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "-----------------------------------"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb33"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb33-1"><a href="#cb33-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(summary.table)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code> region
segment interstitial tumor
Myeloid 20 18</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb35"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb35-1"><a href="#cb35-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Check if the DE results have already been generated</span></span>
<span id="cb35-2"><a href="#cb35-2" aria-hidden="true" tabindex="-1"></a>DE.result.file <span class="ot"><-</span> <span class="fu">grepl</span>(contrast.name, DE.results.files)</span>
<span id="cb35-3"><a href="#cb35-3" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(<span class="fu">sum</span>(DE.result.file) <span class="sc">==</span> <span class="dv">1</span>) {</span>
<span id="cb35-4"><a href="#cb35-4" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb35-5"><a href="#cb35-5" aria-hidden="true" tabindex="-1"></a> <span class="co"># Load the previously generated DE results</span></span>
<span id="cb35-6"><a href="#cb35-6" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> <span class="fu">as.data.frame</span>(<span class="fu">read.csv</span>(<span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>)))</span>
<span id="cb35-7"><a href="#cb35-7" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb35-8"><a href="#cb35-8" aria-hidden="true" tabindex="-1"></a>} <span class="cf">else</span> {</span>
<span id="cb35-9"><a href="#cb35-9" aria-hidden="true" tabindex="-1"></a> <span class="co"># Use the function from the DSPWorkflow package</span></span>
<span id="cb35-10"><a href="#cb35-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb35-11"><a href="#cb35-11" aria-hidden="true" tabindex="-1"></a> <span class="co"># Within slide analysis</span></span>
<span id="cb35-12"><a href="#cb35-12" aria-hidden="true" tabindex="-1"></a> <span class="co"># Listed contrasts are condition, reference </span></span>
<span id="cb35-13"><a href="#cb35-13" aria-hidden="true" tabindex="-1"></a> results.list <span class="ot"><-</span> <span class="fu">diffExpr</span>(<span class="at">object =</span> lmm.input<span class="sc">$</span>subset.object, </span>
<span id="cb35-14"><a href="#cb35-14" aria-hidden="true" tabindex="-1"></a> <span class="at">analysis.type =</span> <span class="st">"Within Groups"</span>, </span>
<span id="cb35-15"><a href="#cb35-15" aria-hidden="true" tabindex="-1"></a> <span class="at">region.col =</span> <span class="st">"region"</span>, </span>
<span id="cb35-16"><a href="#cb35-16" aria-hidden="true" tabindex="-1"></a> <span class="at">regions =</span> <span class="fu">c</span>(<span class="st">"tumor"</span>, <span class="st">"interstitial"</span>), </span>
<span id="cb35-17"><a href="#cb35-17" aria-hidden="true" tabindex="-1"></a> <span class="at">group.col =</span> <span class="st">"segment"</span>, </span>
<span id="cb35-18"><a href="#cb35-18" aria-hidden="true" tabindex="-1"></a> <span class="at">groups =</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>), </span>
<span id="cb35-19"><a href="#cb35-19" aria-hidden="true" tabindex="-1"></a> <span class="at">n.cores =</span> parallel<span class="sc">::</span><span class="fu">detectCores</span>())</span>
<span id="cb35-20"><a href="#cb35-20" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb35-21"><a href="#cb35-21" aria-hidden="true" tabindex="-1"></a> <span class="co"># Create the results df</span></span>
<span id="cb35-22"><a href="#cb35-22" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> results.list<span class="sc">$</span>results</span>
<span id="cb35-23"><a href="#cb35-23" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb35-24"><a href="#cb35-24" aria-hidden="true" tabindex="-1"></a> write.results <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb35-25"><a href="#cb35-25" aria-hidden="true" tabindex="-1"></a> <span class="cf">if</span>(write.results <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb35-26"><a href="#cb35-26" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>))</span>
<span id="cb35-27"><a href="#cb35-27" aria-hidden="true" tabindex="-1"></a> }</span>
<span id="cb35-28"><a href="#cb35-28" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb35-29"><a href="#cb35-29" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>Running Within Group Analysis between Regions
Number of regions in group Myeloid: 2
[1] "Running DE with linear model (no mixed-effect)"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb37"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb37-1"><a href="#cb37-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Adjust column names</span></span>
<span id="cb37-2"><a href="#cb37-2" aria-hidden="true" tabindex="-1"></a>logfc.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"logFC"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb37-3"><a href="#cb37-3" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>logfc <span class="ot"><-</span> results.df[[logfc.column]]</span>
<span id="cb37-4"><a href="#cb37-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-5"><a href="#cb37-5" aria-hidden="true" tabindex="-1"></a>pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"_pval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb37-6"><a href="#cb37-6" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>pval <span class="ot"><-</span> results.df[[pval.column]]</span>
<span id="cb37-7"><a href="#cb37-7" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-8"><a href="#cb37-8" aria-hidden="true" tabindex="-1"></a>adj.pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"adjpval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb37-9"><a href="#cb37-9" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>padj <span class="ot"><-</span> results.df[[adj.pval.column]]</span>
<span id="cb37-10"><a href="#cb37-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-11"><a href="#cb37-11" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>gene <span class="ot"><-</span> results.df<span class="sc">$</span>Gene</span>
<span id="cb37-12"><a href="#cb37-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-13"><a href="#cb37-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Keep only the necessary columns</span></span>
<span id="cb37-14"><a href="#cb37-14" aria-hidden="true" tabindex="-1"></a>results.df <span class="ot"><-</span> results.df <span class="sc">%>%</span> <span class="fu">select</span>(<span class="fu">c</span>(<span class="st">"gene"</span>, </span>
<span id="cb37-15"><a href="#cb37-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"logfc"</span>, </span>
<span id="cb37-16"><a href="#cb37-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"pval"</span>, </span>
<span id="cb37-17"><a href="#cb37-17" aria-hidden="true" tabindex="-1"></a> <span class="st">"padj"</span>))</span>
<span id="cb37-18"><a href="#cb37-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-19"><a href="#cb37-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-20"><a href="#cb37-20" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the results</span></span>
<span id="cb37-21"><a href="#cb37-21" aria-hidden="true" tabindex="-1"></a>export.deg.list <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb37-22"><a href="#cb37-22" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.deg.list <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb37-23"><a href="#cb37-23" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, </span>
<span id="cb37-24"><a href="#cb37-24" aria-hidden="true" tabindex="-1"></a> <span class="at">file =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/DEG_lists/"</span>, contrast.name, <span class="st">"_deg_list.csv"</span>), </span>
<span id="cb37-25"><a href="#cb37-25" aria-hidden="true" tabindex="-1"></a> <span class="at">row.names =</span> <span class="cn">FALSE</span>) </span>
<span id="cb37-26"><a href="#cb37-26" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb37-27"><a href="#cb37-27" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-28"><a href="#cb37-28" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the volcano plot</span></span>
<span id="cb37-29"><a href="#cb37-29" aria-hidden="true" tabindex="-1"></a>volcano.output <span class="ot"><-</span> <span class="fu">make_volcano</span>(<span class="at">lmm.results =</span> results.df, </span>
<span id="cb37-30"><a href="#cb37-30" aria-hidden="true" tabindex="-1"></a> <span class="at">title =</span> contrast.name, </span>
<span id="cb37-31"><a href="#cb37-31" aria-hidden="true" tabindex="-1"></a> <span class="at">legend.title =</span> <span class="st">"Expression in Tumor"</span>, </span>
<span id="cb37-32"><a href="#cb37-32" aria-hidden="true" tabindex="-1"></a> <span class="at">x.axis.title =</span> <span class="st">"Tumor (cond) versus Intersitial (ref) logFC"</span>, </span>
<span id="cb37-33"><a href="#cb37-33" aria-hidden="true" tabindex="-1"></a> <span class="at">fc.limit =</span> <span class="dv">1</span>, </span>
<span id="cb37-34"><a href="#cb37-34" aria-hidden="true" tabindex="-1"></a> <span class="at">pos.label.limit =</span> <span class="dv">1</span>, </span>
<span id="cb37-35"><a href="#cb37-35" aria-hidden="true" tabindex="-1"></a> <span class="at">neg.label.limit =</span> <span class="sc">-</span><span class="fl">2.5</span>)</span>
<span id="cb37-36"><a href="#cb37-36" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-37"><a href="#cb37-37" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the volcano</span></span>
<span id="cb37-38"><a href="#cb37-38" aria-hidden="true" tabindex="-1"></a>export.volcano <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb37-39"><a href="#cb37-39" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.volcano <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb37-40"><a href="#cb37-40" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(<span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/volcano/"</span>, contrast.name, <span class="st">"_volcano_plot.png"</span>), </span>
<span id="cb37-41"><a href="#cb37-41" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb37-42"><a href="#cb37-42" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb37-43"><a href="#cb37-43" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb37-44"><a href="#cb37-44" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb37-45"><a href="#cb37-45" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(volcano.output<span class="sc">$</span>volcano.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast%20Myeloid%20Tumor%20vs%20Myeloid%20Interstitial-1.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb38"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb38-1"><a href="#cb38-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the heatmap</span></span>
<span id="cb38-2"><a href="#cb38-2" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb38-3"><a href="#cb38-3" aria-hidden="true" tabindex="-1"></a><span class="co"># Define new annotations if needed</span></span>
<span id="cb38-4"><a href="#cb38-4" aria-hidden="true" tabindex="-1"></a>anno.colors <span class="ot">=</span> <span class="fu">list</span>(<span class="at">region =</span> <span class="fu">c</span>(<span class="at">tumor =</span> <span class="st">"pink"</span>,</span>
<span id="cb38-5"><a href="#cb38-5" aria-hidden="true" tabindex="-1"></a> <span class="at">interstitial =</span> <span class="st">"seagreen1"</span>, </span>
<span id="cb38-6"><a href="#cb38-6" aria-hidden="true" tabindex="-1"></a> <span class="at">healthyepithelium =</span> <span class="st">"cadetblue1"</span>,</span>
<span id="cb38-7"><a href="#cb38-7" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster =</span> <span class="st">"darkorange"</span>), </span>
<span id="cb38-8"><a href="#cb38-8" aria-hidden="true" tabindex="-1"></a> <span class="at">segment =</span> <span class="fu">c</span>(<span class="at">PanCK =</span> <span class="st">"lightsalmon"</span>, </span>
<span id="cb38-9"><a href="#cb38-9" aria-hidden="true" tabindex="-1"></a> <span class="at">Myeloid =</span> <span class="st">"darkolivegreen1"</span>, </span>
<span id="cb38-10"><a href="#cb38-10" aria-hidden="true" tabindex="-1"></a> <span class="at">other_cells =</span> <span class="st">"lightblue"</span>))</span>
<span id="cb38-11"><a href="#cb38-11" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb38-12"><a href="#cb38-12" aria-hidden="true" tabindex="-1"></a><span class="co"># Subset the annotation just for the heatmap annotations of interest</span></span>
<span id="cb38-13"><a href="#cb38-13" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> lmm.input<span class="sc">$</span>annotation <span class="sc">%>%</span> </span>
<span id="cb38-14"><a href="#cb38-14" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">names</span>(anno.colors))</span>
<span id="cb38-15"><a href="#cb38-15" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb38-16"><a href="#cb38-16" aria-hidden="true" tabindex="-1"></a><span class="co"># Order the annotation</span></span>
<span id="cb38-17"><a href="#cb38-17" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>region), ]</span>
<span id="cb38-18"><a href="#cb38-18" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>segment), ]</span>
<span id="cb38-19"><a href="#cb38-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb38-20"><a href="#cb38-20" aria-hidden="true" tabindex="-1"></a>heatmap.plot <span class="ot"><-</span> <span class="fu">make_heatmap</span>(</span>
<span id="cb38-21"><a href="#cb38-21" aria-hidden="true" tabindex="-1"></a> <span class="at">normalized.log.counts.df =</span> lmm.input<span class="sc">$</span>log.counts, </span>
<span id="cb38-22"><a href="#cb38-22" aria-hidden="true" tabindex="-1"></a> <span class="at">de.results =</span> results.df, </span>
<span id="cb38-23"><a href="#cb38-23" aria-hidden="true" tabindex="-1"></a> <span class="at">top.degs =</span> <span class="cn">TRUE</span>, </span>
<span id="cb38-24"><a href="#cb38-24" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.column =</span> annotation.heatmap,</span>
<span id="cb38-25"><a href="#cb38-25" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.row =</span> <span class="cn">NULL</span>, </span>
<span id="cb38-26"><a href="#cb38-26" aria-hidden="true" tabindex="-1"></a> <span class="at">anno.colors =</span> anno.colors, </span>
<span id="cb38-27"><a href="#cb38-27" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.rows =</span> <span class="cn">TRUE</span>, </span>
<span id="cb38-28"><a href="#cb38-28" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.columns =</span> <span class="cn">TRUE</span>, </span>
<span id="cb38-29"><a href="#cb38-29" aria-hidden="true" tabindex="-1"></a> <span class="at">main.title =</span> contrast.name, </span>
<span id="cb38-30"><a href="#cb38-30" aria-hidden="true" tabindex="-1"></a> <span class="at">row.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb38-31"><a href="#cb38-31" aria-hidden="true" tabindex="-1"></a> <span class="at">column.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb38-32"><a href="#cb38-32" aria-hidden="true" tabindex="-1"></a> <span class="at">show.colnames =</span> <span class="cn">FALSE</span>)</span>
<span id="cb38-33"><a href="#cb38-33" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb38-34"><a href="#cb38-34" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(heatmap.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast%20Myeloid%20Tumor%20vs%20Myeloid%20Interstitial-2.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb39"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb39-1"><a href="#cb39-1" aria-hidden="true" tabindex="-1"></a>export.heatmap <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb39-2"><a href="#cb39-2" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.heatmap <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb39-3"><a href="#cb39-3" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb39-4"><a href="#cb39-4" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(heatmap.plot, </span>
<span id="cb39-5"><a href="#cb39-5" aria-hidden="true" tabindex="-1"></a> <span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/heatmap/"</span>, contrast.name, <span class="st">"_heatmap_plot.png"</span>), </span>
<span id="cb39-6"><a href="#cb39-6" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb39-7"><a href="#cb39-7" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb39-8"><a href="#cb39-8" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb39-9"><a href="#cb39-9" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
</div>
</section>
<section id="de-contrast-myeloid-cluster-vs-myeloid-interstitial" class="level3">
<h3 class="anchored" data-anchor-id="de-contrast-myeloid-cluster-vs-myeloid-interstitial">DE contrast: Myeloid Cluster vs Myeloid Interstitial</h3>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb40"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb40-1"><a href="#cb40-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Setup annotation groups for the contrast</span></span>
<span id="cb40-2"><a href="#cb40-2" aria-hidden="true" tabindex="-1"></a>segment.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>)</span>
<span id="cb40-3"><a href="#cb40-3" aria-hidden="true" tabindex="-1"></a>region.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"cluster"</span>, <span class="st">"interstitial"</span>)</span>
<span id="cb40-4"><a href="#cb40-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-5"><a href="#cb40-5" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-6"><a href="#cb40-6" aria-hidden="true" tabindex="-1"></a><span class="co"># Define the name of the contrast</span></span>
<span id="cb40-7"><a href="#cb40-7" aria-hidden="true" tabindex="-1"></a>contrast.name <span class="ot"><-</span> <span class="fu">paste0</span>(segment.types[[<span class="dv">1</span>]], </span>
<span id="cb40-8"><a href="#cb40-8" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb40-9"><a href="#cb40-9" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">1</span>]], </span>
<span id="cb40-10"><a href="#cb40-10" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb40-11"><a href="#cb40-11" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">2</span>]])</span>
<span id="cb40-12"><a href="#cb40-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-13"><a href="#cb40-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Gather the column names to be used in defining contrasts</span></span>
<span id="cb40-14"><a href="#cb40-14" aria-hidden="true" tabindex="-1"></a>contrast.groups.list <span class="ot"><-</span> <span class="fu">list</span>(</span>
<span id="cb40-15"><a href="#cb40-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"segment"</span> <span class="ot">=</span> segment.types, </span>
<span id="cb40-16"><a href="#cb40-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"region"</span> <span class="ot">=</span> region.types)</span>
<span id="cb40-17"><a href="#cb40-17" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-18"><a href="#cb40-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-19"><a href="#cb40-19" aria-hidden="true" tabindex="-1"></a><span class="co"># Filter data for the chosen annotations</span></span>
<span id="cb40-20"><a href="#cb40-20" aria-hidden="true" tabindex="-1"></a>object <span class="ot"><-</span> normalized.object</span>
<span id="cb40-21"><a href="#cb40-21" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-22"><a href="#cb40-22" aria-hidden="true" tabindex="-1"></a><span class="co"># Generate the counts, annotation, and subset object for lmm</span></span>
<span id="cb40-23"><a href="#cb40-23" aria-hidden="true" tabindex="-1"></a>lmm.input <span class="ot"><-</span> <span class="fu">subset_for_lmm</span>(<span class="at">object =</span> object, </span>
<span id="cb40-24"><a href="#cb40-24" aria-hidden="true" tabindex="-1"></a> <span class="at">subset.list =</span> contrast.groups.list)</span>
<span id="cb40-25"><a href="#cb40-25" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-26"><a href="#cb40-26" aria-hidden="true" tabindex="-1"></a><span class="co"># Create summary table of group counts</span></span>
<span id="cb40-27"><a href="#cb40-27" aria-hidden="true" tabindex="-1"></a>summary.table.df <span class="ot"><-</span> <span class="fu">pData</span>(lmm.input<span class="sc">$</span>subset.object) <span class="sc">%>%</span> </span>
<span id="cb40-28"><a href="#cb40-28" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">c</span>(<span class="fu">names</span>(contrast.groups.list)))</span>
<span id="cb40-29"><a href="#cb40-29" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-30"><a href="#cb40-30" aria-hidden="true" tabindex="-1"></a>summary.table <span class="ot"><-</span> <span class="fu">table</span>(summary.table.df)</span>
<span id="cb40-31"><a href="#cb40-31" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb40-32"><a href="#cb40-32" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"Sample Numbers per Annotation Group"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "Sample Numbers per Annotation Group"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb42"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb42-1"><a href="#cb42-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(<span class="st">"-----------------------------------"</span>)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>[1] "-----------------------------------"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb44"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb44-1"><a href="#cb44-1" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(summary.table)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code> region
segment cluster interstitial
Myeloid 19 20</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb46"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb46-1"><a href="#cb46-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Check if the DE results have already been generated</span></span>
<span id="cb46-2"><a href="#cb46-2" aria-hidden="true" tabindex="-1"></a>DE.result.file <span class="ot"><-</span> <span class="fu">grepl</span>(contrast.name, DE.results.files)</span>
<span id="cb46-3"><a href="#cb46-3" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(<span class="fu">sum</span>(DE.result.file) <span class="sc">==</span> <span class="dv">1</span>) {</span>
<span id="cb46-4"><a href="#cb46-4" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb46-5"><a href="#cb46-5" aria-hidden="true" tabindex="-1"></a> <span class="co"># Load the previously generated DE results</span></span>
<span id="cb46-6"><a href="#cb46-6" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> <span class="fu">as.data.frame</span>(<span class="fu">read.csv</span>(<span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>)))</span>
<span id="cb46-7"><a href="#cb46-7" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb46-8"><a href="#cb46-8" aria-hidden="true" tabindex="-1"></a>} <span class="cf">else</span> {</span>
<span id="cb46-9"><a href="#cb46-9" aria-hidden="true" tabindex="-1"></a> <span class="co"># Use the function from the DSPWorkflow package</span></span>
<span id="cb46-10"><a href="#cb46-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb46-11"><a href="#cb46-11" aria-hidden="true" tabindex="-1"></a> <span class="co"># Within slide analysis</span></span>
<span id="cb46-12"><a href="#cb46-12" aria-hidden="true" tabindex="-1"></a> <span class="co"># Listed contrasts are condition, reference </span></span>
<span id="cb46-13"><a href="#cb46-13" aria-hidden="true" tabindex="-1"></a> results.list <span class="ot"><-</span> <span class="fu">diffExpr</span>(<span class="at">object =</span> lmm.input<span class="sc">$</span>subset.object, </span>
<span id="cb46-14"><a href="#cb46-14" aria-hidden="true" tabindex="-1"></a> <span class="at">analysis.type =</span> <span class="st">"Within Groups"</span>, </span>
<span id="cb46-15"><a href="#cb46-15" aria-hidden="true" tabindex="-1"></a> <span class="at">region.col =</span> <span class="st">"region"</span>, </span>
<span id="cb46-16"><a href="#cb46-16" aria-hidden="true" tabindex="-1"></a> <span class="at">regions =</span> <span class="fu">c</span>(<span class="st">"cluster"</span>, <span class="st">"interstitial"</span>), </span>
<span id="cb46-17"><a href="#cb46-17" aria-hidden="true" tabindex="-1"></a> <span class="at">group.col =</span> <span class="st">"segment"</span>, </span>
<span id="cb46-18"><a href="#cb46-18" aria-hidden="true" tabindex="-1"></a> <span class="at">groups =</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>), </span>
<span id="cb46-19"><a href="#cb46-19" aria-hidden="true" tabindex="-1"></a> <span class="at">n.cores =</span> parallel<span class="sc">::</span><span class="fu">detectCores</span>())</span>
<span id="cb46-20"><a href="#cb46-20" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb46-21"><a href="#cb46-21" aria-hidden="true" tabindex="-1"></a> <span class="co"># Create the results df</span></span>
<span id="cb46-22"><a href="#cb46-22" aria-hidden="true" tabindex="-1"></a> results.df <span class="ot"><-</span> results.list<span class="sc">$</span>results</span>
<span id="cb46-23"><a href="#cb46-23" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb46-24"><a href="#cb46-24" aria-hidden="true" tabindex="-1"></a> write.results <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb46-25"><a href="#cb46-25" aria-hidden="true" tabindex="-1"></a> <span class="cf">if</span>(write.results <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb46-26"><a href="#cb46-26" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/"</span>, contrast.name, <span class="st">"_de.results.csv"</span>))</span>
<span id="cb46-27"><a href="#cb46-27" aria-hidden="true" tabindex="-1"></a> }</span>
<span id="cb46-28"><a href="#cb46-28" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb46-29"><a href="#cb46-29" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output cell-output-stdout">
<pre><code>Running Within Group Analysis between Regions
Number of regions in group Myeloid: 2
[1] "Running DE with linear model (no mixed-effect)"</code></pre>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb48"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb48-1"><a href="#cb48-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Adjust column names</span></span>
<span id="cb48-2"><a href="#cb48-2" aria-hidden="true" tabindex="-1"></a>logfc.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"logFC"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb48-3"><a href="#cb48-3" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>logfc <span class="ot"><-</span> results.df[[logfc.column]]</span>
<span id="cb48-4"><a href="#cb48-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-5"><a href="#cb48-5" aria-hidden="true" tabindex="-1"></a>pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"_pval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb48-6"><a href="#cb48-6" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>pval <span class="ot"><-</span> results.df[[pval.column]]</span>
<span id="cb48-7"><a href="#cb48-7" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-8"><a href="#cb48-8" aria-hidden="true" tabindex="-1"></a>adj.pval.column <span class="ot"><-</span> <span class="fu">colnames</span>(results.df[<span class="fu">grepl</span>(<span class="st">"adjpval"</span>,<span class="fu">colnames</span>(results.df))])</span>
<span id="cb48-9"><a href="#cb48-9" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>padj <span class="ot"><-</span> results.df[[adj.pval.column]]</span>
<span id="cb48-10"><a href="#cb48-10" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-11"><a href="#cb48-11" aria-hidden="true" tabindex="-1"></a>results.df<span class="sc">$</span>gene <span class="ot"><-</span> results.df<span class="sc">$</span>Gene</span>
<span id="cb48-12"><a href="#cb48-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-13"><a href="#cb48-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Keep only the necessary columns</span></span>
<span id="cb48-14"><a href="#cb48-14" aria-hidden="true" tabindex="-1"></a>results.df <span class="ot"><-</span> results.df <span class="sc">%>%</span> <span class="fu">select</span>(<span class="fu">c</span>(<span class="st">"gene"</span>, </span>
<span id="cb48-15"><a href="#cb48-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"logfc"</span>, </span>
<span id="cb48-16"><a href="#cb48-16" aria-hidden="true" tabindex="-1"></a> <span class="st">"pval"</span>, </span>
<span id="cb48-17"><a href="#cb48-17" aria-hidden="true" tabindex="-1"></a> <span class="st">"padj"</span>))</span>
<span id="cb48-18"><a href="#cb48-18" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-19"><a href="#cb48-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-20"><a href="#cb48-20" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the results</span></span>
<span id="cb48-21"><a href="#cb48-21" aria-hidden="true" tabindex="-1"></a>export.deg.list <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb48-22"><a href="#cb48-22" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.deg.list <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb48-23"><a href="#cb48-23" aria-hidden="true" tabindex="-1"></a> <span class="fu">write.csv</span>(results.df, </span>
<span id="cb48-24"><a href="#cb48-24" aria-hidden="true" tabindex="-1"></a> <span class="at">file =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/DEG_lists/"</span>, contrast.name, <span class="st">"_deg_list.csv"</span>), </span>
<span id="cb48-25"><a href="#cb48-25" aria-hidden="true" tabindex="-1"></a> <span class="at">row.names =</span> <span class="cn">FALSE</span>) </span>
<span id="cb48-26"><a href="#cb48-26" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb48-27"><a href="#cb48-27" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-28"><a href="#cb48-28" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the volcano plot</span></span>
<span id="cb48-29"><a href="#cb48-29" aria-hidden="true" tabindex="-1"></a>volcano.output <span class="ot"><-</span> <span class="fu">make_volcano</span>(<span class="at">lmm.results =</span> results.df, </span>
<span id="cb48-30"><a href="#cb48-30" aria-hidden="true" tabindex="-1"></a> <span class="at">title =</span> contrast.name, </span>
<span id="cb48-31"><a href="#cb48-31" aria-hidden="true" tabindex="-1"></a> <span class="at">legend.title =</span> <span class="st">"Expression in Cluster"</span>, </span>
<span id="cb48-32"><a href="#cb48-32" aria-hidden="true" tabindex="-1"></a> <span class="at">x.axis.title =</span> <span class="st">"Cluster (cond) versus Intersitial (ref) logFC"</span>, </span>
<span id="cb48-33"><a href="#cb48-33" aria-hidden="true" tabindex="-1"></a> <span class="at">fc.limit =</span> <span class="dv">1</span>, </span>
<span id="cb48-34"><a href="#cb48-34" aria-hidden="true" tabindex="-1"></a> <span class="at">pos.label.limit =</span> <span class="dv">1</span>, </span>
<span id="cb48-35"><a href="#cb48-35" aria-hidden="true" tabindex="-1"></a> <span class="at">neg.label.limit =</span> <span class="sc">-</span><span class="fl">1.5</span>)</span>
<span id="cb48-36"><a href="#cb48-36" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-37"><a href="#cb48-37" aria-hidden="true" tabindex="-1"></a><span class="co"># Export the volcano</span></span>
<span id="cb48-38"><a href="#cb48-38" aria-hidden="true" tabindex="-1"></a>export.volcano <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb48-39"><a href="#cb48-39" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.volcano <span class="sc">==</span> <span class="cn">TRUE</span>){ </span>
<span id="cb48-40"><a href="#cb48-40" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(<span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/volcano/"</span>, contrast.name, <span class="st">"_volcano_plot.png"</span>), </span>
<span id="cb48-41"><a href="#cb48-41" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb48-42"><a href="#cb48-42" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb48-43"><a href="#cb48-43" aria-hidden="true" tabindex="-1"></a>}</span>
<span id="cb48-44"><a href="#cb48-44" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb48-45"><a href="#cb48-45" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(volcano.output<span class="sc">$</span>volcano.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast%20Myeloid%20Cluster%20vs%20Myeloid%20Interstitial-1.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb49"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb49-1"><a href="#cb49-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Make the heatmap</span></span>
<span id="cb49-2"><a href="#cb49-2" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb49-3"><a href="#cb49-3" aria-hidden="true" tabindex="-1"></a><span class="co"># Define new annotations if needed</span></span>
<span id="cb49-4"><a href="#cb49-4" aria-hidden="true" tabindex="-1"></a>anno.colors <span class="ot">=</span> <span class="fu">list</span>(<span class="at">region =</span> <span class="fu">c</span>(<span class="at">tumor =</span> <span class="st">"pink"</span>,</span>
<span id="cb49-5"><a href="#cb49-5" aria-hidden="true" tabindex="-1"></a> <span class="at">interstitial =</span> <span class="st">"seagreen1"</span>, </span>
<span id="cb49-6"><a href="#cb49-6" aria-hidden="true" tabindex="-1"></a> <span class="at">healthyepithelium =</span> <span class="st">"cadetblue1"</span>,</span>
<span id="cb49-7"><a href="#cb49-7" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster =</span> <span class="st">"darkorange"</span>), </span>
<span id="cb49-8"><a href="#cb49-8" aria-hidden="true" tabindex="-1"></a> <span class="at">segment =</span> <span class="fu">c</span>(<span class="at">PanCK =</span> <span class="st">"lightsalmon"</span>, </span>
<span id="cb49-9"><a href="#cb49-9" aria-hidden="true" tabindex="-1"></a> <span class="at">Myeloid =</span> <span class="st">"darkolivegreen1"</span>, </span>
<span id="cb49-10"><a href="#cb49-10" aria-hidden="true" tabindex="-1"></a> <span class="at">other_cells =</span> <span class="st">"lightblue"</span>))</span>
<span id="cb49-11"><a href="#cb49-11" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb49-12"><a href="#cb49-12" aria-hidden="true" tabindex="-1"></a><span class="co"># Subset the annotation just for the heatmap annotations of interest</span></span>
<span id="cb49-13"><a href="#cb49-13" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> lmm.input<span class="sc">$</span>annotation <span class="sc">%>%</span> </span>
<span id="cb49-14"><a href="#cb49-14" aria-hidden="true" tabindex="-1"></a> <span class="fu">select</span>(<span class="fu">names</span>(anno.colors))</span>
<span id="cb49-15"><a href="#cb49-15" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb49-16"><a href="#cb49-16" aria-hidden="true" tabindex="-1"></a><span class="co"># Order the annotation</span></span>
<span id="cb49-17"><a href="#cb49-17" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>region), ]</span>
<span id="cb49-18"><a href="#cb49-18" aria-hidden="true" tabindex="-1"></a>annotation.heatmap <span class="ot"><-</span> annotation.heatmap[<span class="fu">order</span>(annotation.heatmap<span class="sc">$</span>segment), ]</span>
<span id="cb49-19"><a href="#cb49-19" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb49-20"><a href="#cb49-20" aria-hidden="true" tabindex="-1"></a>heatmap.plot <span class="ot"><-</span> <span class="fu">make_heatmap</span>(</span>
<span id="cb49-21"><a href="#cb49-21" aria-hidden="true" tabindex="-1"></a> <span class="at">normalized.log.counts.df =</span> lmm.input<span class="sc">$</span>log.counts, </span>
<span id="cb49-22"><a href="#cb49-22" aria-hidden="true" tabindex="-1"></a> <span class="at">de.results =</span> results.df, </span>
<span id="cb49-23"><a href="#cb49-23" aria-hidden="true" tabindex="-1"></a> <span class="at">top.degs =</span> <span class="cn">TRUE</span>, </span>
<span id="cb49-24"><a href="#cb49-24" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.column =</span> annotation.heatmap,</span>
<span id="cb49-25"><a href="#cb49-25" aria-hidden="true" tabindex="-1"></a> <span class="at">annotation.row =</span> <span class="cn">NULL</span>, </span>
<span id="cb49-26"><a href="#cb49-26" aria-hidden="true" tabindex="-1"></a> <span class="at">anno.colors =</span> anno.colors, </span>
<span id="cb49-27"><a href="#cb49-27" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.rows =</span> <span class="cn">TRUE</span>, </span>
<span id="cb49-28"><a href="#cb49-28" aria-hidden="true" tabindex="-1"></a> <span class="at">cluster.columns =</span> <span class="cn">TRUE</span>, </span>
<span id="cb49-29"><a href="#cb49-29" aria-hidden="true" tabindex="-1"></a> <span class="at">main.title =</span> contrast.name, </span>
<span id="cb49-30"><a href="#cb49-30" aria-hidden="true" tabindex="-1"></a> <span class="at">row.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb49-31"><a href="#cb49-31" aria-hidden="true" tabindex="-1"></a> <span class="at">column.gaps =</span> <span class="cn">NULL</span>, </span>
<span id="cb49-32"><a href="#cb49-32" aria-hidden="true" tabindex="-1"></a> <span class="at">show.colnames =</span> <span class="cn">FALSE</span>)</span>
<span id="cb49-33"><a href="#cb49-33" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb49-34"><a href="#cb49-34" aria-hidden="true" tabindex="-1"></a><span class="fu">print</span>(heatmap.plot)</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
<div class="cell-output-display">
<p><img src="CPTR9_DE_Analysis_files/figure-html/DE%20contrast%20Myeloid%20Cluster%20vs%20Myeloid%20Interstitial-2.png" class="img-fluid" width="672"></p>
</div>
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb50"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb50-1"><a href="#cb50-1" aria-hidden="true" tabindex="-1"></a>export.heatmap <span class="ot"><-</span> <span class="cn">FALSE</span></span>
<span id="cb50-2"><a href="#cb50-2" aria-hidden="true" tabindex="-1"></a><span class="cf">if</span>(export.heatmap <span class="sc">==</span> <span class="cn">TRUE</span>){</span>
<span id="cb50-3"><a href="#cb50-3" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb50-4"><a href="#cb50-4" aria-hidden="true" tabindex="-1"></a> <span class="fu">ggsave</span>(heatmap.plot, </span>
<span id="cb50-5"><a href="#cb50-5" aria-hidden="true" tabindex="-1"></a> <span class="at">filename =</span> <span class="fu">paste0</span>(results.folder, run.folder, <span class="st">"DE/heatmap/"</span>, contrast.name, <span class="st">"_heatmap_plot.png"</span>), </span>
<span id="cb50-6"><a href="#cb50-6" aria-hidden="true" tabindex="-1"></a> <span class="at">width =</span> <span class="dv">14</span>, </span>
<span id="cb50-7"><a href="#cb50-7" aria-hidden="true" tabindex="-1"></a> <span class="at">height =</span> <span class="dv">10</span>)</span>
<span id="cb50-8"><a href="#cb50-8" aria-hidden="true" tabindex="-1"></a> </span>
<span id="cb50-9"><a href="#cb50-9" aria-hidden="true" tabindex="-1"></a>}</span></code><button title="Copy to Clipboard" class="code-copy-button"><i class="bi"></i></button></pre></div>
</details>
</div>
</section>
<section id="de-contrast-myeloid-cluster-vs-myeloid-tumor" class="level3">
<h3 class="anchored" data-anchor-id="de-contrast-myeloid-cluster-vs-myeloid-tumor">DE contrast: Myeloid Cluster vs Myeloid Tumor</h3>
<div class="cell">
<details>
<summary>Code</summary>
<div class="sourceCode cell-code" id="cb51"><pre class="sourceCode r code-with-copy"><code class="sourceCode r"><span id="cb51-1"><a href="#cb51-1" aria-hidden="true" tabindex="-1"></a><span class="co"># Setup annotation groups for the contrast</span></span>
<span id="cb51-2"><a href="#cb51-2" aria-hidden="true" tabindex="-1"></a>segment.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"Myeloid"</span>)</span>
<span id="cb51-3"><a href="#cb51-3" aria-hidden="true" tabindex="-1"></a>region.types <span class="ot"><-</span> <span class="fu">c</span>(<span class="st">"cluster"</span>, <span class="st">"tumor"</span>)</span>
<span id="cb51-4"><a href="#cb51-4" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb51-5"><a href="#cb51-5" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb51-6"><a href="#cb51-6" aria-hidden="true" tabindex="-1"></a><span class="co"># Define the name of the contrast</span></span>
<span id="cb51-7"><a href="#cb51-7" aria-hidden="true" tabindex="-1"></a>contrast.name <span class="ot"><-</span> <span class="fu">paste0</span>(segment.types[[<span class="dv">1</span>]], </span>
<span id="cb51-8"><a href="#cb51-8" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb51-9"><a href="#cb51-9" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">1</span>]], </span>
<span id="cb51-10"><a href="#cb51-10" aria-hidden="true" tabindex="-1"></a> <span class="st">"_"</span>, </span>
<span id="cb51-11"><a href="#cb51-11" aria-hidden="true" tabindex="-1"></a> region.types[[<span class="dv">2</span>]])</span>
<span id="cb51-12"><a href="#cb51-12" aria-hidden="true" tabindex="-1"></a></span>
<span id="cb51-13"><a href="#cb51-13" aria-hidden="true" tabindex="-1"></a><span class="co"># Gather the column names to be used in defining contrasts</span></span>
<span id="cb51-14"><a href="#cb51-14" aria-hidden="true" tabindex="-1"></a>contrast.groups.list <span class="ot"><-</span> <span class="fu">list</span>(</span>
<span id="cb51-15"><a href="#cb51-15" aria-hidden="true" tabindex="-1"></a> <span class="st">"segment"</span> <span class="ot">=</span> segment.types, </span>