4b. HISAT

You will now try the splice aware aligner HISAT (~20 minutes). This exercise can be completed automatically by running /home/ubuntu/hisat/doHISAT.sh. Pre-computed results can be found in /home/ubuntu/hisat/cached.

First, index the references for use by Hisat.

Commands to type:

$ cd ~/genome
$ hisat-build chr20.fa chr20.hisat

Screen output:

Settings:
  Output files: "chr20.hisat.*.bt2"
  Line rate: 6 (line is 64 bytes)
     ...
Total time for backward call to driver() for mirror index: 00:00:40

File output:

chr20.fa	chr20.hisat.1.bt2	chr20.hisat.2.bt2	chr20.hisat.3.bt2	chr20.hisat.4.bt2	chr20.hisat.5.bt2	chr20.hisat.6.bt2	chr20.hisat.rev.1.bt2	chr20.hisat.rev.2.bt2	chr20.hisat.rev.5.bt2	chr20.hisat.rev.6.bt2

You will now start alignment for hisat.

Commands to type:

$ cd ~/hisat
$ hisat -p 16 -x ../genome/chr20.hisat -U ../rawdata/SRR1153470_1.fastq -S hisat.sam

Here is some information about what each argument does:

-p: splits up the alignment into 16 jobs

-x: instructs the aligner where to look for the reference file

-U: instructs the aligner where to look for the input file

-S: instructs the aligner what to name the output file 

Again, convert the .sam file to an indexed .bam file so that you can visualize mapped reads later in the exercise.

Commands to type:

$ samtools view -bSF4 hisat.sam | samtools sort - hisat.sorted
$ samtools index hisat.sorted.bam