samtools returns error – cigar and query sequence are of different length even though cigar and query sequence are the same length

samtools returns error – cigar and query sequence are of different length even though cigar and query sequence are the same length

1

I have written a program in python that processes and outputs mapped results as either a bam or sam file. It works fine until I have indels within the sequence. when I try to process the result file using samtools, it returns the following error:

samtools [e::sam_parse1] cigar and query sequence are of different length"

…even though the cigar and query sequence are of the same length (see below sample sam lines which returned the error).

mapped_read1    0   chr1    1379078 60  17M1D58M    =   1379078 0   CTATCAATTTTTTGTTTTTTGTTGTTGTTGTTTTTTAAGGATTGTGGGCCGGGTACAGTGACTCACGCCTGTAATC    AAAEEEEEE/EEAEE/EEEEEEEEEEEEEEEEEEEEEEE6<EAE/EEE<AEEEEEEAE/EEEEEEEEAEEEEEEEE    MD:Z:13T3^G58   XG:Z:1  NH:i:1  NM:Z:1  XM:Z:1  XN:Z:0  XO:Z:1  AS:Z:-13    YT:Z:UU
mapped_read2    99  chr1    1654630 60  34M2D24M    =   1654630 211 ATTCTACTTTATTTGTGCAAAATCTTTTTTTTCCTTTTTTTTTTTAGAGGCGGGGTCTTG    EEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEAEEEEEEEEEEEEE    MD:Z:34^T^T24   XG:Z:2  NH:i:1  NM:Z:0  XM:Z:0  XN:Z:0  XO:Z:1  AS:Z:-11    YT:Z:0
mapped_read3    0   chr1    1684108 60  24M1D43M    =   1684108 0   CACTCAGCAGTCGTGAGACGCTTGCCCAATTGCCTTTTCTTTTAAAGGCCCCTGCCATTTCCCAAACC    AAAEEEEEEEEEEEEAEEEEEEEEEEEEEEEEE<EEEEEEEEEEEEEEEE<EEEEEEEEEAEEEEEEE    MD:Z:24^C37A5   XG:Z:1  NH:i:1  NM:Z:1  XM:Z:1  XN:Z:0  XO:Z:1  AS:Z:-11    YT:Z:UU
mapped_read 0   chr1    1889614 60  47M1D72M    =   1889614 0   ACCAGCCTGGTTAACAGGATCTCTTTTACAAAGAGATCCCATCTCTTAAAAAAAAAAAAAAAAAAGGCTTCCATGAAGATGAATTAAGCAAACAAAAGCTGATACTGCTTCTGCATGCTA    AAAAEEE/EEEEEEEAEEEEEEEEAEEEEEEE/EEAEEEAEEEAEEEEEEEEEEEEEEEEEEEE/EEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEAEEEEEEEEEEEEEEEEEEEEEE    MD:Z:47^A72 XG:Z:1  NH:i:1  NM:Z:0  XM:Z:0  XN:Z:0  XO:Z:1  AS:Z:-8 YT:Z:UU
mapped_read4    0   chr1    2286565 60  58M1D10M    =   2286565 0   GATTTGAACCCCAAATATAATAATTAAAGTGAGTGTGCATTTTATTCTTACCACATCAGGCAAACGCTG   EEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEE/EEEEE   MD:Z:31C26^G10  XG:Z:1  NH:i:1  NM:Z:1  XM:Z:1  XN:Z:0  XO:Z:1  AS:Z:-13    YT:Z:UU
mapped_read5    0   chr1    2946932 60  9M1D59M =   2946932 0   ACAAAACCTGGCCCACCCTGGTGAGGGGAGTTCTAGCCTCTCCCAGGCCCCCATGGCACGATCTAAGTG   AAAEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEE   MD:Z:9^G25C33   XG:Z:1  NH:i:1  NM:Z:1  XM:Z:1  XN:Z:0  XO:Z:1  AS:Z:-13    YT:Z:UU
mapped_read6    0   chr1    4479047 60  42M2D57M    =   4479047 0   GTGTAAAAGTGTTCCTTTTCACCACATCCATGCCAACGTCTATATTTTTTTTTTGGATTTTATATTAATTGGCCATTCTTGCAGGAGTAAGGTGGTATCTC   AAAEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEEE//EEEEEEEEEEAAEEEEEEEEEEEEEEEEEEEEEEEEAEEEEEEEE   MD:Z:42^T^T57   XG:Z:2  NH:i:1  NM:Z:0  XM:Z:0  XN:Z:0  XO:Z:1  AS:Z:-11    YT:Z:UU

This result bam/sam file was generated using the pysam module in python. Any help would be appreciated. I have tried using samtools 0.1.18, 1.2, 1.6, 1.13, all with the same error.


sam


samtools


bam

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