title: RNA seq of ND_R1_rep1
design: Total RNA was extracted using the ReliaPrep RNA Miniprep Systems (Promega) according to the manufacturers instructions. 1000 ng of total RNA was used to synthesis cDNA using M-MLV Reverse Transcriptase, RNase H Minus, Point Mutant (Promega) according to the manufacturers instructions. NGS short reads were trimmed using fastp(Chen et al., 2018) and processed using Kalliso, an RNAseq quantification program that uses a pseudoalignment method of  assigning reads to genomic locations in lieu of a more costly traditional alignment(Bray et al., 2016). The human transcriptome, version GRCh38.p12(Schneider et al., 2017), was indexed, the paired, trimmed reads assigned to transcripts, and read counts converted to transcripts per million (TPM) by Kallisto. TPMs from transcripts originating from the same gene were aggregated and relative expression levels were computed as the log2 fold change relative to the matched wild type using an in-house script (available upon request). RPKM values were computed from TPMs using the median transcript length per gene.
library_name: 2