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6 hours ago
mk
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300
According to the STAR docs, soft-clipping using the --clip[3p,5p]Nbases option automatically clips the corresponding end of the read during alignment, so that we don't try to map adapters.
In the following example, I clipped the 5-prime 10X adapter,bc,umi by setting STAR ... --clip5pNbases 41 0....
Just looking at the R1 FWD reads shows many reads whose CIGAR has a clip (S) value is less than 41, which seems to ignore the directive above to clip (at least) 41 bases:
(/nfs/turbo/[...]/shared3012) [user@gl1527 FCA_gut8015057_S1_L001_bigwig]$ samtools view -f 99 /tmpssd/user/test_chr1_min40.bam|head -n 4
A00708:13:HTC7MDSXX:4:1364:26720:35462 99 chr1 14467 255 39S62M = 14642 240 ACGCAGCGTCGAATCTTCTGTGCTGGTTTCTTATATGGGGGCGCAGGCTGGGTGGAGCCGTCCCCCCATGGAGCACAGGCAGACAGAAGTCCCCGCCCCAG FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF NH:i:1 HI:i:1 nM:i:0 AS:i:125 CR:Z:ACGCAGCGTCGAATCT UR:Z:TCTGTGCTGG GX:Z:ENSG00000310526.1 GN:Z:WASH7P sS:Z:ACGCAGCGTCGAATCTTCTGTGCTGGTTTCTTATATGGGGGCGCAGGCTGGGTGGAGCCGTCCCCCCATGGAGCACAGGCAGACAGAAGTCCCCGCCCCAG sQ:Z:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF,FF:FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF sM:i:0 CB:Z:ACGCAGCGTCGAATCT UB:Z:TCTGTGCTGG
A00708:13:HTC7MDSXX:4:2668:2311:21997 99 chr1 17470 255 39S62M = 17703 477 CCCTCCTCATCAGGGGCACAGCGTGCACTGGGGGGTCCCAGGCCTCCCGAGCCGAGCCACCCGTCACCCCCTGGCTCCTGGCCTATGTGCTGTACCTGTGT ,FFFFF,FFFFFFFFFFFFFFFFFFF,,,F:,,,,F,,,FFFFFFFFFFF:FFFFFFFFFFF:,FFFFFFFFFFFFFF:FFFFFFFF:FFFFFFFFFFFF: NH:i:1 HI:i:1 nM:i:4 AS:i:126 CR:Z:CCCTCCTCATCAGGGG UR:Z:CACAGCGTGC GX:Z:ENSG00000310526.1 GN:Z:WASH7P sS:Z:CCCTCCTCATCAGGGGCACAGCGTGCACTGGGGGGTCCCAGGCCTCCCGAGCCGAGCCACCCGTCACCCCCTGGCTCCTGGCCTATGTGCTGTACCTGTGT sQ:Z:,FFFFF,FFFFFFFFFFFFFFFFFFF,,,F:,,,,F,,,FFFFFFFFFFF:FFFFFFFFFFF:,FFFFFFFFFFFFFF:FFFFFFFF:FFFFFFFFFFFF: sM:i:-1 CB:Z:- UB:Z:-
A00708:13:HTC7MDSXX:4:2668:1750:23469 99 chr1 17470 255 39S62M = 17703 477 CCCTCCTCATCAGGGGCACAGCGTGCACTGTGGGGTCCCAGGCCTCCCGAGCCGAGCCACCCGTCACCCCCTGGCTCCTGGCCTATGTGCTGTACCTGTGT FFFFFFFFFFFFFFFFFFFFFFFFFF,,,::,,,,F,,,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF NH:i:1 HI:i:1 nM:i:3 AS:i:128 CR:Z:CCCTCCTCATCAGGGG UR:Z:CACAGCGTGC GX:Z:ENSG00000310526.1 GN:Z:WASH7P sS:Z:CCCTCCTCATCAGGGGCACAGCGTGCACTGTGGGGTCCCAGGCCTCCCGAGCCGAGCCACCCGTCACCCCCTGGCTCCTGGCCTATGTGCTGTACCTGTGT sQ:Z:FFFFFFFFFFFFFFFFFFFFFFFFFF,,,::,,,,F,,,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF sM:i:-1 CB:Z:- UB:Z:-
A00708:13:HTC7MDSXX:4:2253:19913:14904 99 chr1 17470 255 39S62M = 17656 459 CCCTCCTCATCAGGGGCACAGCGTGCACTGTGGGGTCCCAGGCCTCCCGAGCCGAGCCACCCGTCACCCCCTGGCTCCTGGCCTATGTGCTGTACCTGTGT FFFFFFFFFFFFFFFFFFFFFFFFFF,,FF,,,,,:,,,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF NH:i:1 HI:i:1 nM:i:0 AS:i:163 CR:Z:CCCTCCTCATCAGGGG UR:Z:CACAGCGTGC GX:Z:ENSG00000310526.1 GN:Z:WASH7P sS:Z:CCCTCCTCATCAGGGGCACAGCGTGCACTGTGGGGTCCCAGGCCTCCCGAGCCGAGCCACCCGTCACCCCCTGGCTCCTGGCCTATGTGCTGTACCTGTGT sQ:Z:FFFFFFFFFFFFFFFFFFFFFFFFFF,,FF,,,,,:,,,FFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFFF sM:i:-1 CB:Z:- UB:Z:-
