RNA-seq pipeline includes Quality Control, rRNA filtering, Genome Alignment using HISAT2, STAR and Tophat2, and estimating gene and isoform expression levels by RSEM and featureCounts.

Steps:

1. For Quality Control, we use FastQC to create qc outputs. There are optional read quality filtering (trimmomatic), read quality trimming (trimmomatic), adapter removal (cutadapt) processes available.
2. Bowtie2/Bowtie/STAR is used to count or filter out common RNAs (eg. rRNA, miRNA, tRNA, piRNA etc.).
3. RSEM is used to align RNA-Seq reads to a reference transcripts and estimates gene and isoform expression levels.
4. HISAT2, STAR and Tophat2 is used to align RNA-Seq reads to a genome. Optional estimation of gene and isoform expression levels could be done by featureCounts.
5. Genome-wide Bam analysis is done by RseQC, Picard.
6. Optionally you can create Integrative Genomics Viewer (IGV) and Genome Browser Files (TDF and Bigwig, respectively)

Program Versions:

• FastQC v0.11.8
• Star v2.6.1
• Hisat2 v2.1.0
• Picard v2.18.27
• Rseqc v2.6.2
• Samtools v1.3
• Subread v1.6.4
• Multiqc v1.7
• Tophat v2.1.1
• RSEM v1.3.1
• Bowtie2 v2.3.5
• Bowtie v1.2.2
• Trimmomatic v0.39
• Igvtools v2.5.3
• Bedtools v2.27.1
• Fastx_toolkit v0.0.14
• Ucsc-wigToBigWig v366
• Pdfbox-App v2.0.0

Run through DolphinNext User Interface:

To start using the dolphinnext/rnaseq pipeline please go to DolphinNext Web page and click run button.

Run through Command Line:

To install and start using the dolphinnext/rnaseq pipeline by using command line, please follow these steps: Installation.