I am wondering about to use a BAM file output as a input to make a alignment to others FASTA reference genome. I have been reading how to get a FASTA formand from my BAM file but I am not sure if the reads depth could affect my alignment. If that is possible, How could I do that?
Hi Lissa Cruz Saavedra ,
Your question is a bit confusing. But, if you want to use reads in bam file (for alignment or whatever) I recommend you to use bedtools to extract fastq reads from bam file as described here: https://bedtools.readthedocs.io/en/latest/content/tools/bamtofastq.html.
Bam file contains your reads (exactly as your originals, but in a different format), PLUS the coordinates where they were aligned and the score of the alignment. So, I do not what you mean by; "the read depth could affect your alignment".
In addition, I recommend you not to use fasta sequences for alignment if you can use fastq.
Best regards.
Thanks dear Francisco
I am sorry if I wasn't absolutely clear. I was thinking to use fasta because I would like to include some fasta from reference samples in my analysis. In that case will I need to change the fasta format to fastq as well?.
No, in that case, extract the sequences from Bam in fasta, you can not change fasta to fastq unless you have quality scores. Remember that the main difference between fasta and fastq is that fastq have quality scores for each base sequenced. Sometimes you can change fasta to fastq assigning arbitrary quality scores, but it is used just for exploratory purposes because it is a biased analysis.
Besides, if you mix sequencing reads with fasta (from reference samples), of course, RDC will affect your analysis. In such case I recommend you to reduce redundancy (from BAM) before mix samples.
Thanks a lot. I am sorry but I don't have too much experience about work with bam files, just one question more, how could I reduce the redundancy in my bam files before to change the format to fasta?.
Probably the most easy way is using CD-HIT, but you need to extract the sequences from bam first. It is used for clustering sequences based on its similitude (you must use the representative sequence from each cluster), so, in this way you will count each genomic fragment (or whatever reference) only once.
Before extract sequences from BAM, I recommend you read about SAM flags, and avoid multi-mapping reads.
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