📄
Guide to RNA-seq Analysis
  • Introduction
  • Preparations
    • Work environment
    • Softwares and databases
    • Obtain sequencing data
    • R packages
  • Raw read processing
    • QC & trimming
    • Mapping & quantification
      • Alignment-free method
      • Alignment-based method
  • Differential expression analysis in R
    • Building a TxDb object
    • About tximport
    • Convert Salmon output to Sleuth-compatible format
    • Differential gene expression (DGE) analysis using DESeq2
      • DGE analysis with Salmon/Kallisto input
      • DGE analysis with STAR input
      • DGE analysis with STAR + RSEM input
    • Differential transcript expression (DTE) analysis using DESeq2
      • DTE analysis with Salmon/Kallisto input
      • DTE analysis with STAR + RSEM input
    • DGE and DTE analysis of Salmon/Kallisto inputs using Sleuth
    • Differential transcript usage (DTU) analysis
      • DTU analysis using DRIMSeq
      • DTU analysis using DEXSeq
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  • Environment
  • Directories

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  1. Preparations

Work environment

Environment

  • CPU: Intel® Xeon® E5-2623 v3 (Quad-Core; 8 Threads)

  • Memory: 64 GB

  • Ubuntu 18.04.4 LTS

  • R version 3.6.2 (2019-12-12)

CPU usage is limited to 6 threads and memory to 32 GB where applicable

Directories

Path

Purpose

/home/USER/tools

Software/tools

/home/USER/db/refanno

Reference genome, annotation and index

/home/USER/SSAPs

RNA-seq exercise on sessile serrated adenomas/polyps dataset

/home/USER/SSAPs/fastqs

FASTQ files

/home/USER/SSAPs/trimmed

Trimmed FASTQ files

/home/USER/SSAPs/salmon

Output from Salmon

/home/USER/SSAPs/star

Output from STAR

/home/USER/SSAPs/salmon_star

Output from STAR+Salmon

/home/USER/SSAPs/rsem_star

Output from STAR+RSEM

/home/USER/SSAPs/analysis

R analysis (DGE, DTE, and DTU)

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Last updated 5 years ago

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