[caddymob]

How are you seeing the header then that you posted previously?

[urchgene]

@ caddymob...........................not sure i understood your question but

i copied and pasted the @HD VN:1.0 SO:coordinate into a txt file header.txt as you advised.

but since that did not work, i tried this again

samtools view -H stub_header.sam > header.txt

but the header file was empty because there was an error report in the run.

[caddymob]

Ok, you have a sam, not a bam? Then just edit the sam header directly. Then samtools import the sam to make it a bam. Should be good to go.

[adarob]

@jeffhsu3,

Ok. I spent a few hours on this and figured it out. It is a problem with mis-parsing the unmapped hits. It only affects the BAM parser, so for now you can convert to SAM and everything should work fine. The next version (later this week) will include the fix for BAM. Thanks for bringing this to our attention.

-Adam

[brachysclereid]

Cufflinks Error: sort order of reads in BAMs must be the same

Hi, I am having a similar problem to that mentioned in this thread. I am using TopHat v1.1.4 to create the BAMs with from bowtie indexes. My cufflinks version is v0.9.3.

$ cufflinks -G annotation.gtf ./s_1/accepted_hits.bam
cufflinks: /usr/lib64/libz.so.1: no version information available (required by cufflinks)
Error: sort order of reads in BAMs must be the same

Based on previous posts I tried reverting from the BAM to SAM, but got the same result. Would I need to convert BAM to SAM and then sort? If the GTF is the problem how should one sort this file? Reading through the previous post it was unclear to me what the ultimate solution was. Reordering the GTF or putting new headers on the BAM?

[torfnase]

Hi, I have the very same problem. I am using TopHat v.1.2.0 to create BAMs. My cufflink version is 1.0.3
cufflinks ../../tophat/M2/accepted_hits_Sample_M2-2_wt_300.bam
Error: sort order of reads in BAMs must be the same
I tried to revert the BAM to SAM but still the same error.
Furthermore, I exchanged the header of the BAM file. But also in this instance the same error.
What else could I try?
Thanks

[adarob]

torfnase, can you send just the header to [email protected]?

[songsy]

adarob, I have a similar question. I first use tophat to get the bam file and then run cufflinks with --GTF command, which goes wrong with message like:
You are using Cufflinks v2.0.2, which is the most recent release.
Error: sort order of reads in BAMs must be the same

The order of the GFF file is the same with the header of bam files:

9 ensembl chromosome 1 156750706 . . . ID=9;Name=chromosome:AGPv2:9:1:156750706:1
1 ensembl chromosome 1 301354135 . . . ID=1;Name=chromosome:AGPv2:1:1:301354135:1
4 ensembl chromosome 1 241473504 . . . ID=4;Name=chromosome:AGPv2:4:1:241473504:1
5 ensembl chromosome 1 217872852 . . . ID=5;Name=chromosome:AGPv2:5:1:217872852:1
2 ensembl chromosome 1 237068873 . . . ID=2;Name=chromosome:AGPv2:2:1:237068873:1
3 ensembl chromosome 1 232140174 . . . ID=3;Name=chromosome:AGPv2:3:1:232140174:1
6 ensembl chromosome 1 169174353 . . . ID=6;Name=chromosome:AGPv2:6:1:169174353:1
8 ensembl chromosome 1 175793759 . . . ID=8;Name=chromosome:AGPv2:8:1:175793759:1
7 ensembl chromosome 1 176764762 . . . ID=7;Name=chromosome:AGPv2:7:1:176764762:1
10 ensembl chromosome 1 150189435 . . . ID=10;Name=chromosome:AGPv2:10:1:150189435:1
UNKNOWN ensembl chromosome 1 7140151 . . . ID=UNKNOWN;Name=chromosome:AGPv2:UNKNOWN:1:7140151:1
Pt ensembl chromosome 1 140384 . . . ID=Pt;Name=chromosome:AGPv2:Pt:1:140384:1
Mt ensembl chromosome 1 569630 . . . ID=Mt;Name=chromosome:AGPv2:Mt:1:569630:1

And the header of bam file is:
@HD VN:1.0 SO:coordinate
@SQ SN:chromosome:AGPv2:9:1:156750706:1 LN:156750706
@SQ SN:chromosome:AGPv2:1:1:301354135:1 LN:301354135
@SQ SN:chromosome:AGPv2:4:1:241473504:1 LN:241473504
@SQ SN:chromosome:AGPv2:5:1:217872852:1 LN:217872852
@SQ SN:chromosome:AGPv2:2:1:237068873:1 LN:237068873
@SQ SN:chromosome:AGPv2:3:1:232140174:1 LN:232140174
@SQ SN:chromosome:AGPv2:6:1:169174353:1 LN:169174353
@SQ SN:chromosome:AGPv2:8:1:175793759:1 LN:175793759
@SQ SN:chromosome:AGPv2:7:1:176764762:1 LN:176764762
@SQ SN:chromosome:AGPv2:10:1:150189435:1 LN:150189435
@SQ SN:chromosome:AGPv2:UNKNOWN:1:7140151:1 LN:7140151
@SQ SN:chromosome:AGPv2:Pt:1:140384:1 LN:140384
@SQ SN:chromosome:AGPv2:Mt:1:569630:1 LN:569630

I don't know what's wrong??? Thank you very much!!!

@lindymcb,

Are you using a GTF annotation? If so, it needs to be in the same order as your SAM file OR you must use a SAM header.

[adarob]

Colons (":") cannot be used as separators in the reference names.