GATK DepthofCoverage Error

[bwubb]

Im trying to use GATK DepthofCoverage, but Im getting an error which I do not understand how to resolve.

My command is:

java -Xmx4g -jar ~/GenomeAnalysisTK-1.3/GenomeAnalysisTK.jar \
-T DepthOfCoverage \
-R ~/b37_genomes/human_g1k_v37.fasta \
-I PE.merged.sorted.recal.bam \
-L ~/BED/exons.b37.bed \
-o test.GATK.out

But then I get:

##### ERROR MESSAGE: File associated with name ~/BED/exons.b37.bed is malformed: Interval file could not be parsed in any supported format. caused by BED files must be parsed through Tribble; parsing them as intervals through the GATK engine is no longer supported
##### ERROR ------------------------------------------------------------------------------------------

The GATK wiki does not have many examples of what can be used. This is Whole Exome sequencing data, so naturally I want to determine the coverage over all exon intervals.

I have no clue what this Tribble thing is, and how can they not support BED format....

Thanks.

[Heisman]

Could you post the header lines of your file and then the first few intervals as well?

[bwubb]

Wow. Sorry I missed the reply to this thread. Thank for responding.

Here are the first few headers:

@HD	VN:1.0	GO:none	SO:coordinate
@SQ	SN:1	LN:249250621
@SQ	SN:2	LN:243199373
@SQ	SN:3	LN:198022430
@SQ	SN:4	LN:191154276
@SQ	SN:5	LN:180915260
@SQ	SN:6	LN:171115067
@SQ	SN:7	LN:159138663
@SQ	SN:8	LN:146364022
@SQ	SN:9	LN:141213431

Is this sufficient, or did you want the alignments as well? Thanks again for the help, this is still an issue for me.

[Heisman]

It indicates the bed file is the problem: could you post maybe the first 50 lines of that?

[bwubb]

1	14467	14587
1	14639	14883
1	14943	15064
1	15671	15990
1	16591	16719
1	16750	17074
1	17178	17420
1	17443	18108
1	18203	18448
1	19049	19170
1	20603	20723
1	24448	24915
1	29267	29389
1	30275	30431
1	35095	35215
1	35245	35366
1	35668	35788
1	62983	63703
1	69069	70029
1	112710	112830
1	120754	120966
1	129018	129259
1	133356	133597
1	135235	135355
1	135688	136176
1	137366	137726
1	173754	173874
1	228233	228711
1	259027	259147
1	267075	267287
1	326408	326768
1	327177	327665
1	327998	328118
1	329752	329993
1	334092	334333
1	342357	342569
1	350498	350618
1	367647	368608
1	470971	471330
1	621084	622045
1	639075	639195
1	647124	647336
1	655375	655616
1	659720	659961
1	661601	661721
1	662054	662542
1	662854	663214
1	709552	709672
1	717360	717480
1	721338	721640

It is b37 format, but I have been consistent in its use. This is not my design file bed, but rather "all" exons. I wouldnt think this is the issue though. If there is no overlap and shouldnt it just report zero coverage?

[Heisman]

So I use the "1.0.2885" version of GATK for DepthOfCoverage and I use interval files that look like this:

@HD     VN:1.0  SO:unsorted
@SQ     SN:chr1 LN:249250621    UR:file:hg19.fa       M5:1b22b98cdeb4a9304cb5d48026a85128
@SQ     SN:chr2 LN:243199373    UR:file:hg19.fa       M5:a0d9851da00400dec1098a9255ac712e
@SQ     SN:chr3 LN:198022430    UR:file:hg19.fa       M5:641e4338fa8d52a5b781bd2a2c08d3c3
@SQ     SN:chr4 LN:191154276    UR:file:hg19.fa       M5:23dccd106897542ad87d2765d28a19a1
@SQ     SN:chr5 LN:180915260    UR:file:hg19.fa       M5:0740173db9ffd264d728f32784845cd7
@SQ     SN:chr6 LN:171115067    UR:file:hg19.fa       M5:1d3a93a248d92a729ee764823acbbc6b
@SQ     SN:chr7 LN:159138663    UR:file:hg19.fa       M5:618366e953d6aaad97dbe4777c29375e
@SQ     SN:chr8 LN:146364022    UR:file:hg19.fa       M5:96f514a9929e410c6651697bded59aec
@SQ     SN:chr9 LN:141213431    UR:file:hg19.fa       M5:3e273117f15e0a400f01055d9f393768
@SQ     SN:chr10        LN:135534747    UR:file:hg19.fa       M5:988c28e000e84c26d552359af1ea2e1d
@SQ     SN:chr11        LN:135006516    UR:file:hg19.fa       M5:98c59049a2df285c76ffb1c6db8f8b96
@SQ     SN:chr12        LN:133851895    UR:file:hg19.fa       M5:51851ac0e1a115847ad36449b0015864
@SQ     SN:chr13        LN:115169878    UR:file:hg19.fa       M5:283f8d7892baa81b510a015719ca7b0b
@SQ     SN:chr14        LN:107349540    UR:file:hg19.fa       M5:98f3cae32b2a2e9524bc19813927542e
@SQ     SN:chr15        LN:102531392    UR:file:hg19.fa       M5:e5645a794a8238215b2cd77acb95a078
@SQ     SN:chr16        LN:90354753     UR:file:hg19.fa       M5:fc9b1a7b42b97a864f56b348b06095e6
@SQ     SN:chr17        LN:81195210     UR:file:hg19.fa       M5:351f64d4f4f9ddd45b35336ad97aa6de
@SQ     SN:chr18        LN:78077248     UR:file:hg19.fa       M5:b15d4b2d29dde9d3e4f93d1d0f2cbc9c
@SQ     SN:chr19        LN:59128983     UR:file:hg19.fa       M5:1aacd71f30db8e561810913e0b72636d
@SQ     SN:chr20        LN:63025520     UR:file:hg19.fa       M5:0dec9660ec1efaaf33281c0d5ea2560f
@SQ     SN:chr21        LN:48129895     UR:file:hg19.fa       M5:2979a6085bfe28e3ad6f552f361ed74d
@SQ     SN:chr22        LN:51304566     UR:file:hg19.fa       M5:a718acaa6135fdca8357d5bfe94211dd
@SQ     SN:chrM LN:16571        UR:file:hg19.fa       M5:d2ed829b8a1628d16cbeee88e88e39eb
@SQ     SN:chrX LN:155270560    UR:file:hg19.fa       M5:7e0e2e580297b7764e31dbc80c2540dd
@SQ     SN:chrY LN:59373566     UR:file:hg19.fa       M5:1e86411d73e6f00a10590f976be01623
chr1    2985721 2985854 +       target_1
chr1    3102668 3103058 +       target_2
chr1    3160630 3160721 +       target_3

with the header created as described here: http://www.broadinstitute.org/gsa/wi...ference_genome

Maybe try something like that?