HISAT2-index-align
The DE Quick Start tutorial provides an introduction to basic DE functionality and navigation.
Rationale and background:
HISAT2: Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown
Mihaela Pertea,Daehwan Kim, Geo M Pertea, Jeffrey T Leek, Steven L Salzberg
Nature Protocols 11,1650–1667(2016)doi:10.1038/nprot.2016.095
HISAT2 software (http://ccb.jhu.edu/software/hisat2 or http://github.com/infphilo/hisat2, version 2.0.1 or later)
Pre-Requisites
- A CyVerse account. (Register for an CyVerse account here - user.cyverse.org)
- Mandatory arguments
- Output folder name
- Input file reference genome sequence in fasta format
- FASTQ Files (Read 1) : Input reads 1 files of paired end data or reads of single end data
- FASTQ Files (Read 2) : Input reads 2 files of paired end data or leave this field empty for single end data
- Fragment Library Type: specify the format of the library- more details(http://sailfish.readthedocs.io/en/master/library_type.html)
- File type: Enter whether the library is paired end or single end
- Optional arguments:
- Trim bases from 5' end of read:Trim bases from 5' (left) end of each read before alignment
- Trim bases from 3' end of read: Trim bases from 3' (right) end of each read before alignment
Phred quality score: encoding for quality score
- Minimum intron length:Sets minimum intron length
- maximum intron length:Sets maximum intron length
- Report alignments tailored for transcript assemblers including StringTie:With this option, HISAT2 requires longer anchor lengths for de novo discovery of splice sites. This leads to fewer alignments with short-anchors, which helps transcript assemblers improve significantly in computational and memory usage.
- Report alignments tailored for transcript assemblers including StringTie:With this option, HISAT2 requires longer anchor lengths for de novo discovery of splice sites. This leads to fewer alignments with short-anchors, which helps transcript assemblers improve significantly in computational and memory usage.
- minimum fragment length for valid paired-end alignments:The minimum fragment length for valid paired-end alignments.
- maximum fragment length for valid paired-end alignments:
The following test data are provided for testing HISAT2 in here - /iplant/home/shared/iplantcollaborative/example_data/tophat2-PE( We will use a similar data as used for tophat2-PE):
- left_reads- SRR946914_fastq_1.fastq,SRR946916_fastq_1.fastq
- right_reads-SRR946914_fastq_2.fastq, SRR946916_fastq_2.fastq
- reference-NC_010473.fa
Results
Successful execution of the HISAT2-index-align assessment pipeline will create a directory named out. The directory will contain bam and bai files for each sample. This can be used for further downstream analysis and visualization purpose:
output
SRR946914_fastq_1.sorted.bam
SRR946914_fastq_1.sorted.bam.bai
SRR946916_fastq_1.sorted.bam
SRR946916_fastq_1.sorted.bam.bai