User: ATPoint
ATPoint • 720
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Posts by ATPoint
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... The reference fasta uses only the numbers to name the chromosomes, so 1,2,3 instead of chr1, chr2, chr3.
You can change that in awk, as [previously][1] suggested::
awk '/^>/{print ">chr" ++i; next}{print}' < foo.fasta
[1]: https://www.biostars.org/p/53212/ ...
written 13 hours ago by
ATPoint • 720
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... The situation you describe would lead to a low mapping quality during the alignment, probably even 0. Good practice is to discard these reads categorically because, as you say, one cannot distinguish properly. I typically remove duplicates after all the filtering steps. For standard assays like ATAC ...
written 15 hours ago by
ATPoint • 720
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... Ok, it seems that you have the TruSeq Ribo adapter. You can doublecheck with [this file from Illumina][1], containing the common adapters. In any case, it is not Nextera. Retrim with the proper adapter sequence and the issue should be solved.
[1]: https://support.illumina.com/content/dam/illumin ...
written 1 day ago by
ATPoint • 720
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... No, as in paired-end, the insert size, which defines the fragment, are solely defined by the 5' ends of the respective fwd and rev reads. Have a look at [this figure][1], you can see that no matter where the 3' ends are (so the arrow heads), the 5' ends are unaffected by this, and so is the insert s ...
written 1 day ago by
ATPoint • 720
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... There are two things that you (I think) are mixing up.
1) trimming is done on the fastq files, but rmdup works on aligned data
2) duplicates are defined by the 5' ends of the paired-end data, while trimming takes away bases from the 3' end, so no worries, trimming will not affect duplicate removal ...
written 1 day ago by
ATPoint • 720
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Comment:
C: weird insert size post trimming
... Also be sure that the adapter file is correct.
CTGTCTCTTATACACATCTCCGAGCCCACGAGAC is what you have to trim for in case that the Nextera adapter was used for library prep (on both strands, same adapter sequence). ...
written 1 day ago by
ATPoint • 720
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Comment:
C: NGS compute infrastructure
... Make sure to have sufficient I/O capacity to really make full use of CPU and RAM. The best cluster makes no sense if the I/O bottleneck kills all the performance and permits to use multithreading effectively. ...
written 3 days ago by
ATPoint • 720
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Comment:
C: Low Alignment Rate for TT2 Cells
... The bowtie-index is the same for all the different cell lines experiments? Did you manually blast some of the unmapped reads, maybe a contamination in the TT2 stock? ...
written 3 days ago by
ATPoint • 720
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... I think the 256 flag only means that it is not a primary alignment. This can then be a chimeric one that has been tagged as 256 due to the -M option, but also a multimapping read. If reads multimap, then one is randomly chosen to be primary, all others get flagged as secondary. That means SA:Z is on ...
written 4 days ago by
ATPoint • 720
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3
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Comment:
C: weird insert size post trimming
... Please give the full command for cutadapt. I saw this kind of odd shape before, but it turned out that adapters were improperly trimmed. What are the Nextera sequences that you trimmed for? ...
written 4 days ago by
ATPoint • 720
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