Missed alignments STAR vs Bowtie2
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Nel
Apr 5, 2018, 3:28:01 PM4/5/18
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Hi,
I'm using STAR on metatranscriptomic data from a decaying plant. I've also tried Bowtie2 and among all alignments only found by Bowtie2 some of them don't seem so terrible to me (it's actually from my plant according to BLAST results)
Do you have any idea why STAR missed them ? I've run STAR with default parameters, but with an annotation file.
Besides, since I don't have a fully assembled genome for reference, are there some parameters to tweak ?
Thank you for your help!
Nel
I'm using STAR on metatranscriptomic data from a decaying plant. I've also tried Bowtie2 and among all alignments only found by Bowtie2 some of them don't seem so terrible to me (it's actually from my plant according to BLAST results)
Besides, since I don't have a fully assembled genome for reference, are there some parameters to tweak ?
Thank you for your help!
Nel
Alexander Dobin
Apr 7, 2018, 4:28:46 PM4/7/18
to rna-star
Hi Nel,
the alignment did not paste well, please paste it as plain text.
Also, please for this read please try to map read1 and read2 separately with STAR, and post the results.
Cheers
Alex
On Thursday, April 5, 2018 at 3:28:01 PM UTC-4, Nel wrote:
Hi,
I'm using STAR on metatranscriptomic data from a decaying plant. I've also tried Bowtie2 and among all alignments only found by Bowtie2 some of them don't seem so terrible to me (it's actually from my plant according to BLAST results)
Here is an example :
Read1 99 scaffold346 52047 44 150M = 52110 213 GTTGATTGTGAAGATATCTCCATATGGCTGATAGTGATGTGGGTTTGAGATTGGCTCGGCTGATATTCCTGCGTAGTCCGTTGTAAAGACTATAGGTTTCCCATCCGGGCTGAAATACGGGTGGTTGCACCGCCCGCCCGGCCCGCTCTG AS:i:295 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:95C54 YS:i:300 YT:Z:CP Read2 147 scaffold346 52110 44 150M = 52047 -213 TATTCCTGCGTAGTCCGTTGTAAAGACTATAGCTTTCCCATCCGGGCTGAAATACGGGTGGTTGCACCGCCCGCCCGGCCCGCTCTGGACGAGTTTTCTCAAACCGGTTCCGTTCGGGTGGATCAGGTAAATCGCGAAGCTCCCACTTCC AS:i:300 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:150 YS:i:295 YT:Z:CP
Nel
Apr 9, 2018, 8:37:11 AM4/9/18
to rna-...@googlegroups.com
Hi Alex,
Actually I had an issue with SAMtools (while extracting mapped reads some of them were not print (sick!)).
I've found another pair of reads that was only mapped by Bowtie2, even if the alignment is not as good, it's still seem these sequences belong to the plant.
With separate STAR analyses, the log says 100% uniquely mapped reads :
R1 0 scaffold251 102428 255 150M * 0 0 CCGGTCAAGGTCTTCACAAAGATCTGCATACCTCCCCTCAGCCTGAGGACCAGGTGAAGGGTTGACTCCTTTTGAATATTGTAATCGGCCAAAGTACGGCCGTCCTCCAGCTGCTTTCCGGCGAAGATCAGCCTCTGCTGGTCCGGGGGG NH:i:1HI:i:1 AS:i:148 nM:i:0
R2 0 scaffold121 202418 255 150M * 0 0 GGCTAAGATCCAGGACAAGGAAGGGATCCCTCCGGATCAGCAGAGGTTGATCTTCGCCGGAAAGCAGCTCGAGGACGGCCGTACCTTGGCCGATTACAATATTCAGAAGGAGTCAACCCTTCATCTTGTCCTCAGGCTCAGGGGAGGTAT NH:i:1HI:i:1 AS:i:142 nM:i:3
Bowtie2 results :
R1 99 scaffold251 102428 14 150M = 102455 177 CCGGTCAAGGTCTTCACAAAGATCTGCATACCTCCCCTCAGCCTGAGGACCAGGTGAAGGGTTGACTCCTTTTGAATATTGTAATCGGCCAAAGTACGGCCGTCCTCCAGCTGCTTTCCGGCGAAGATCAGCCTCTGCTGGTCCGGGGGG AS:i:300 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:150 YS:i:230 YT:Z:CP
R2 147 scaffold251 102455 14 146M4S = 102428 -177 ATACCTCCCCTGAGCCTGAGGACAAGATGAAGGGTTGACTCCTTCTGAATATTGTAATCGGCCAAGGTACGGCCGTCCTCGAGCTGCTTTCCGGCGAAGATCAACCTCTGCTGATCCGGAGGGATCCCTTCCTTGTCCTGGATCTTAGCC AS:i:230 XS:i:285 XN:i:0 XM:i:10 XO:i:0 XG:i:0 NM:i:10 MD:Z:11C11C2G17T20A14C22G9G5G17T8 YS:i:300 YT:Z:CP
Thank you for your help !
Nel
Actually I had an issue with SAMtools (while extracting mapped reads some of them were not print (sick!)).
I've found another pair of reads that was only mapped by Bowtie2, even if the alignment is not as good, it's still seem these sequences belong to the plant.
With separate STAR analyses, the log says 100% uniquely mapped reads :
R1 0 scaffold251 102428 255 150M * 0 0 CCGGTCAAGGTCTTCACAAAGATCTGCATACCTCCCCTCAGCCTGAGGACCAGGTGAAGGGTTGACTCCTTTTGAATATTGTAATCGGCCAAAGTACGGCCGTCCTCCAGCTGCTTTCCGGCGAAGATCAGCCTCTGCTGGTCCGGGGGG NH:i:1HI:i:1 AS:i:148 nM:i:0
R2 0 scaffold121 202418 255 150M * 0 0 GGCTAAGATCCAGGACAAGGAAGGGATCCCTCCGGATCAGCAGAGGTTGATCTTCGCCGGAAAGCAGCTCGAGGACGGCCGTACCTTGGCCGATTACAATATTCAGAAGGAGTCAACCCTTCATCTTGTCCTCAGGCTCAGGGGAGGTAT NH:i:1HI:i:1 AS:i:142 nM:i:3
Bowtie2 results :
R1 99 scaffold251 102428 14 150M = 102455 177 CCGGTCAAGGTCTTCACAAAGATCTGCATACCTCCCCTCAGCCTGAGGACCAGGTGAAGGGTTGACTCCTTTTGAATATTGTAATCGGCCAAAGTACGGCCGTCCTCCAGCTGCTTTCCGGCGAAGATCAGCCTCTGCTGGTCCGGGGGG AS:i:300 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:150 YS:i:230 YT:Z:CP
R2 147 scaffold251 102455 14 146M4S = 102428 -177 ATACCTCCCCTGAGCCTGAGGACAAGATGAAGGGTTGACTCCTTCTGAATATTGTAATCGGCCAAGGTACGGCCGTCCTCGAGCTGCTTTCCGGCGAAGATCAACCTCTGCTGATCCGGAGGGATCCCTTCCTTGTCCTGGATCTTAGCC AS:i:230 XS:i:285 XN:i:0 XM:i:10 XO:i:0 XG:i:0 NM:i:10 MD:Z:11C11C2G17T20A14C22G9G5G17T8 YS:i:300 YT:Z:CP
Thank you for your help !
Nel
Alexander Dobin
Apr 9, 2018, 5:03:44 PM4/9/18
to rna-star
Hi Nel,
the R2 alignment is quite bad actually, it has 10 mismatches, so STAR probably did not find any good seeds for R2 near R1 alignment.
Cheers
Alex
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