from CRAM to fastq

Question

[machaalani29]

hello, so I have some WGS data, and I was given the cram and cram.crai; I want to change them to fastq I am using samtools:

samtools fastq -O -c 7 C-7507T.cram > C-7507T.fastq.gz

The cram file is 32G The fastq file is 583G

is that difference in size normal?

[GenoMax]

Possibly. You appear to have written the data to a single file (in case it was paired-end to begin with). Check to make sure secondary alignments did not result in duplicate reads entries.

[machaalani29]

one of the issues is that I am not very sure how the sequencing was done, but I think it was single end. any suggestions hoot do that?

[ATpoint]

I am 99.99 sure that your fastq file is not compressed. What is file C-7507T.fastq.gz? I do not think samtools autodetects suffix if you send to stdout like that.

[machaalani29]

the option -c is to compress the file

[ATpoint]

Yes, but I still think that with this syntax you get an uncompressed file, because samtools does not detect the gz suffix. Just take a head of the fastq file and see whether it is text or binary.

Answer 1

[Pierre Lindenbaum]

if the CRAM/BAM is paire-end, it must be sorted on query name using samtools collate

https://x.com/lh3lh3/status/1132756684789768202

Command line to remap a position sorted bam:

samtools collate -uOn128 old-pos-srt.bam tmpxyz | samtools fastq - | bwa mem -pt16 ref.fa - | samtools sort --threads=4 -m4G -o new-pos-srt.bam