# Phasing problem using "samtools phase"

so I'm having a problem using samtools phase. I'm trying to phase my .bam archives that where generated by aligning my samples with my reference genome. Following the documentation I tried the command:

samtools phase -A my_referenceGenome.fasta my_sample_toBePhased.bam

The -A flag stands for dropping reads with ambiguous phase. The error I get is: [W::sam_read1] Parse error at line 1. I generated .bam files with and without the SAM header but the error persists.

I feel like I am missing something basic but that isn't clearly stated anywere. Does the my_referenceGenome.fasta archive needs to be previously phased? If so, how can I do it?

Thank you so much!

This is what I see in my samtools phase (v1.10)

Usage:   samtools phase [options] <in.bam>


samtools phase only takes a bam file as an input. But you are specifying a .fasta file as an input. If you want to specify the reference genome, you should be using the argument "--reference" as described here

      --reference FILE
Reference sequence FASTA FILE [null]

• Oh, you're right! I also tried that but it didn't work. I managed to make it work and I'll add an answer shortly. But before that I want to ask a little question: in my pipeline I also remove pcr duplicates with the fixmate/markdup steps. Is it better to phase the reads before or after removing the duplicates? Apr 23 '20 at 13:34
• I would recommend removing duplicates first. Apr 23 '20 at 18:05