Hi,
I am attempting to use CollectRnaSeqMetrics from picard tools on my BAM files but I am not being successful with it for some reason. Below is the code that I am using:
The problem is the next line:
So it says that 0 genes are being loaded, but then it goes on to process the lines in the BAM file and the resulting output says all the reads are intergenic as it is not finding any genes. I have tried it with several other refFlat files, but cant seem to get the program to load any genes with any of the files.
I downloaded the refFlat file from UCSC and changed the chromosome names so they match the BAM file
refFlat file:
BAM file:
Just for reference, the fastq files were paired end and aligned using tophat2 using GRCh38 index.
If anyone can help me with this, it would be much appreciated,
Thanks
I am attempting to use CollectRnaSeqMetrics from picard tools on my BAM files but I am not being successful with it for some reason. Below is the code that I am using:
PHP Code:
java -jar /local/software/picard-tools/1.97/jarlib/CollectRnaSeqMetrics.jar REF_FLAT=refFlat.txt STRAND=SECOND_READ_TRANSCRIPTION_STRAND I=Pre_1.bam O=RnaSeqMetrics_1.txt
PHP Code:
INFO 2016-01-08 08:54:50 CollectRnaSeqMetrics Loaded 0 genes.
I downloaded the refFlat file from UCSC and changed the chromosome names so they match the BAM file
refFlat file:
PHP Code:
HSD3B2 NM_000198 1 + 119415119 119423039 119415419 119422620 4 "119415119,119415330,119419417,119421808," "119415202,119415561,119419582,119423039,"
ANKRD65 NM_001243536 1 - 1418419 1421444 1419541 1421005 3 "1418419,1420815,1421132," "1419549,1421005,1421444,"
ANKRD65 NM_001243535 1 - 1418419 1421444 1419233 1421005 3 "1418419,1420796,1421132," "1419549,1421005,1421444,"
PDIK1L NM_001243533 1 + 26111776 26125548 26114308 26122577 4 "26111776,26112459,26114291,26121836," "26111915,26112650,26114593,26125548,"
PDIK1L NM_001243532 1 + 26111164 26125548 26114308 26122577 3 "26111164,26114291,26121836," "26111420,26114593,26125548,"
SLC25A24 NM_013386 1 - 108134721 108200358 108136652 108200138 10 "108134721,108139057,108143542,108148278,108154982,108157461,108161181,108181940,108185827,108199955," "108136837,108139208,108143710,108148386,108155135,108157620,108161293,108182028,108185954,108200358,"
SLC27A3 NM_001317929 1 + 153775291 153780157 153775356 153780002 10 "153775291,153776517,153777061,153777760,153778160,153778462,153778686,153779113,153779342,153779930," "153776164,153776727,153777220,153777885,153778355,153778553,153778885,153779211,153779473,153780157,"
ALG6 NM_013339 1 + 63367589 63438562 63370977 63437020 15 "63367589,63370770,63396512,63402253,63404452,63406316,63407061,63411145,63411925,63414060,63415872,63419369,63428732,63428927,63436822," "63367687,63371059,63396597,63402343,63404541,63406399,63407126,63411331,63412061,63414146,63415957,63419440,63428801,63429126,63438562,"
TPM3 NR_103461 1 - 154155303 154183249 154183249 154183249 8 "154155303,154170399,154170648,154172028,154172907,154173083,154176114,154182987," "154157721,154170465,154170711,154172104,154172978,154173201,154176248,154183249,"
NR1I3 NM_001077481 1 - 161229665 161238210 161229796 161236565 8 "161229665,161230812,161231328,161232806,161233168,161235846,161236458,161238040," "161229926,161230933,161231474,161232946,161233338,161235977,161236598,161238210,"
PHP Code:
HWI-ST1041:151:C7BJEACXX:1:1101:1127:17612 83 8 125231917 50 35M = 125231901 -51 TCAGTATATGCTGGCATACTGAATAAATAAATTAA 4D@:4>>:9?@@BBA>>@>>AAB;>5>5;3>1;5, AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:35 YT:Z:UU NH:i:1 XS:A:+
HWI-ST1041:151:C7BJEACXX:1:1101:1127:17612 163 8 125231901 50 51M = 125231917 51 TGGTACATGGTAGTAGTCAGTATATGCTGGCATACTGAATAAATAAATTAA 0-3:-;)=>@:99=>)9<4;@8@4@????<?<?48;>>>??8=?<.===<> AS:i:-2 XN:i:0 XM:i:1 XO:i:0 XG:i:0 NM:i:1 MD:Z:15T35 YT:Z:UU NH:i:1 XS:A:+
HWI-ST1041:151:C7BJEACXX:1:1101:1128:82805 83 5 81259354 50 43M = 81259277 -120 CTTTCTCTATCTACAATATAAAAATAAGAAAAAAATTAAGTTT AB=A?0???14=ABBAA@:BBBBBABB?ACB=CCA<+A<2++1 AS:i:0 XN:i:0 XM:i:0 XO:i:0 XG:i:0 NM:i:0 MD:Z:43 YT:Z:UU NH:i:1 XS:A:+
HWI-ST1041:151:C7BJEACXX:1:1101:1128:82805 163 5 81259277 50 100M = 81259354 120 CCTGCTGGGCCAGGATCAGCTCAGATGGGACTGCTTTGTGGAGGAAGAAAACATCACTGCCCATTCCTTAACCCTTACTTTCTCTATCTACAATATAAAA 0;9?(==???>9>?8??>?<?@?<>?@):.=?????=>??9)9=-39.7<>5>=>??>;8>;>>?@@@>).8=?;>==>????=>>>??==5;:3==:;; AS:i:-4 XN:i:0 XM:i:2 XO:i:0 XG:i:0 NM:i:2 MD:Z:4A22C72 YT:Z:UU NH:i:1 XS:A:+
If anyone can help me with this, it would be much appreciated,
Thanks