Bam file of RNA-seq

[wangli]

Hi, forks

When I import my RNA-seq in BAM format to the CLC genomic workbench, it automatically turns out to be two files with different file name. I thougth these two files are the same. But when I did reads-mapping of the two files separately, I found that total reads of two files are different, also the percentage of mapped reads varied with the same parameter settings.

Very confused with these two files. Could anyone help to explain it? Thanks!

[jnfass]

Are there different read groups, perhaps two different samples in the BAM? How was it produced (tool, command example, etc.)? Maybe you could show examples of SAM lines from each of the two files ...

[wangli]

The bam file is generated by sequencing company. And they are indicated as one sample.
Could you tell me how to show SAM lines from the two files?
Thanks!

[jnfass]

If you're comfortable working on the command line (linux/unix, Mac OS X terminal?) and have samtools installed, use 'samtools view' ...
If not, can you get helpful details about the aligned reads by hovering over features in CLC?

[wangli]

I get samtools installed and it is a huge file.
which command can be used to see the file page by page?
I tried less | samtools view myfile.bam.
It didnot work.

[kopi-o]

Maybe CLC Bio partitions the files into paired and non-paired files. Could this be waht happened? (It does this with FASTQ files, after trimming and discarding bad reads)

samtools view myfile.bam |less

should work for you.

[wangli]

Thank you, kopi-o!

Both of them are pointing to paired files. One is *** [no read group](paired), the other is ***(paired).

[kopi-o]

OK, so maybe your BAM file contains some alignments for reads tagged with a read group (@RG) and some without a read group. It could be a merged file of alignments from two different instrument runs, for instance. Could that be it?