One of our goals is to visualize our RNA-seq data using a genome browser.
To prepare alignment data for upload to IGV, the RNAseq_analyzer script simply performed some file format conversions.
file.sam –> file.bam This compresses the .sam file into a binary file
file.bam –> file_sort.bam This sorts the .bam file by chromosome location
file_sort.bam.bai This makes a simple indexing reference file for the sorted .bam file
file.bam –> file.bx This creates a bigwig file
These files can be downloaded from summit using Cyberduck. They can then be opened with IGV.
First, we'll go through how to do this using IGV.
C. elegans (ce11)
EGO01_sort.bam.bai. You can do this by navigating to these files in the file structure, right clicking on them, and selecting Download.
You can not compare the heights of any genome browser plots until you have normalized their heights. This is because the height of each plot is proportional to the number of fragments that were sequenced over all. Until you normalize, .bam files from samples that were sequenced to 50 million reads will look twice as tall as those sequenced to 25 million reads.
To normalize the samples:
To remove autoscaling