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Based on pre-alignment QA/QC, we need to trim low quality bases from the 3' end of reads. 

• Select Click the Unaligned reads data node
• Select Trim bases from the Click Pre-alignment tools section of tools in the task menu
• Click Trim bases (Figure 1) 
  
  

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By default, Trim bases removes bases starting at the 3' end of reads and continuing until it reaches finds a base pair call with a Phred score of equal to or greater than 20. Hover over any Image Removed to get additional information about a specific option. 

• Set Trim based on to Quality score
• Set End min quality level (Phred) to 20 
• Set Trim from end to 3-prime (right end)
• Leave Advanced options set to default
• Select Finish (Figure 2)
  

35 (Figure 2). 

• Click Finish to run Trim bases with default settings
  

The Trim bases task will generate a new data node, Trimmed reads. While tasks have been queued or are in progress they have a lighter color. Any output nodes that the task will generate, such as the  (Figure 3). We can view the task report for Trim bases by double-clicking either the Trim bases task node or the Trimmed reads data node from the Trim bases task, are also displayed in a lighter color until the task completes. Once the task begins running, a progress bar is displayed on the task node (Figure 3).  or choosing Task report from the task menu. 

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• Double-click the Trimmed reads data node Select Task report from to open the task menureport

The report shows the percentage of trimmed reads and reads removed in a spreadsheet (Figure 4) and a graph two graphs (Figure 54).  

The average quality score for each base call at each position for each sample is also shown as a graph (Figure 6).

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The results are fairly consistent across samples with ~65% ~2% of reads untrimmed, ~30% ~86% trimmed, and ~5% ~12% removed for each.   The average quality score for each sample is increased with higher average quality scores at the 3' ends. 

• Click RNA-Seq 5-AZA to return to the Analyses tab

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