Short Intro to 

shRNA Single-Cell

RNA-seq

Sample Information

• 96 Samples
• Single Lane ?-  274.44 M reads

Processing Steps

- RNA-seq

• IGenomes UCSC mm9
• Tophat 2 Unstranded with ERCC spikes
• FeatureCounts only paired ---Rsubread 
• DESEQ2 - Standard analysis
  • using RUVseq to get spike in adjustments.
  • rlog transform for PCA.

Processing Steps

shRNA-seq

• pRLL_shRNA_Rep_library in csv
• Full shRNA FASTA contigs built in R
• Duplicate names removed.
• Indexed and aligned using bwa.
• Properly paired and unique mapping hits retained
• Counts for shRNAs in samples established from mapped BAMS.

shRNA

• Alignments to shRNA inspected in IGV.
• shRNA alignments realigned to genome.
• Cross contamination of reads in shRNA to target gene and reverse.

Distribution of shRNAs

RNA-seq

Associated to MTOR -ve PC2 loading

Associated to Cdkn1a/Camk2a