Mark duplicates after fastq trimming?
Hello!
I am fairly new to bioinformatics and have a question about using Picard Mark Duplicates after trimming Illumina sequence reads.
(I could not find the exact answer to my question, but I apologize if this a repeat question. Also, I originally posted this question as a reply to a really old thread (linked below), but thought perhaps it was more appropriate/helpful to start a new thread. I could not figure out how to delete my original post.)
I am using Trimmomatic to trim low quality bases and adapters from paired end whole genome DNA re-sequencing data (fastq files). I am aligning this data to a reference and will ultimately use the GATK to call snps.
My question is: Does anyone know how Picard Mark Duplicates knows which bases were trimmed from a read? If you trim bases from the 5' end of the read, how does Picard know that? Hoes does it know what the original 5' mapping position would have been, and thus which reads are potentially pcr duplicates? See link and quoted comment from user xguo below.
Thank you!
Amanda
Originally Posted by xguo on this old thread:
" The following is what I found from samtools-help list:
"Essentially what Picard does (for pairs; single-end data is also handled) is to find the 5' coordinates and mapping orientations of each read pair. When doing this it takes into account all clipping that has taking place as well as any gaps or jumps in the alignment. You can thus think of it as determining "if all the bases from the read were aligned, where would the 5' most base
have been aligned". It then matches all read pairs that have identical 5' coordinates and orientations and marks as duplicates all but the "best" pair. "Best" is defined as the read pair having the highest sum of base qualities as bases with Q >= 15." "
I am fairly new to bioinformatics and have a question about using Picard Mark Duplicates after trimming Illumina sequence reads.
(I could not find the exact answer to my question, but I apologize if this a repeat question. Also, I originally posted this question as a reply to a really old thread (linked below), but thought perhaps it was more appropriate/helpful to start a new thread. I could not figure out how to delete my original post.)
I am using Trimmomatic to trim low quality bases and adapters from paired end whole genome DNA re-sequencing data (fastq files). I am aligning this data to a reference and will ultimately use the GATK to call snps.
My question is: Does anyone know how Picard Mark Duplicates knows which bases were trimmed from a read? If you trim bases from the 5' end of the read, how does Picard know that? Hoes does it know what the original 5' mapping position would have been, and thus which reads are potentially pcr duplicates? See link and quoted comment from user xguo below.
Thank you!
Amanda
Originally Posted by xguo on this old thread:
" The following is what I found from samtools-help list:
"Essentially what Picard does (for pairs; single-end data is also handled) is to find the 5' coordinates and mapping orientations of each read pair. When doing this it takes into account all clipping that has taking place as well as any gaps or jumps in the alignment. You can thus think of it as determining "if all the bases from the read were aligned, where would the 5' most base
have been aligned". It then matches all read pairs that have identical 5' coordinates and orientations and marks as duplicates all but the "best" pair. "Best" is defined as the read pair having the highest sum of base qualities as bases with Q >= 15." "
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