RNA-seq workshop November 2018, University of Otago

Here you can find most of the scripts that were used during the RNA-seq workshop.

Workshop outline

• Overview of sequencing technologies

• Presentation of RNA-seq libraries

• What is a Fastq file?

• How to QC a fastq ?

• How to trim a sequence file?

  • Quality trimming
  • Adaptor trimming

• How to align the sequencing reads on a reference using HISAT2

  • creating a reference sequence
  • aligning reads on a reference

• Defining alignment format

• Introduction to SAMtools

• How to get a count matrix

  • What is an annotation file
  • How to use featureCount

• LUNCH BREAK

• Differential Gene Expression analysis

  • Using linear model with Bayesian models: Limma
  • Using a negative binomial distribution approach with DESeq2

• Discussion of p-values and hypothesis testing

• Multiple test correction

• Discussion on RNA-seq unit: CPM, R/FPKM and TPM

• How to plot RNA-seq results?

• DISCUSSION AND FOLLOW-UP