Cufflinks -G Problems

[ayfchang]

I am using cufflinks 0.9.3.Linux_x86_64

I have encountered problems running cufflinks -G option as in

cufflinks -p 16 -G mm_ncbi_37_v60.gtf -o ./outputdir accepted_hits.bam

I am using a filtered version of GTF based on mouse ncbi37 ensembl v60 GTF as reference

18 protein_coding exon 3281686 3281767 . - . gene_id "ENSMUSG00000063889"; transcript_id "ENSMUST00000140332"; exon_number "1"; gene_name "Crem"; transcript_name "Crem-025";
18 protein_coding exon 3273420 3273576 . - . gene_id "ENSMUSG00000063889"; transcript_id "ENSMUST00000140332"; exon_number "2"; gene_name "Crem"; transcript_name "Crem-025";
18 protein_coding exon 3266955 3267730 . - . gene_id "ENSMUSG00000063889"; transcript_id "ENSMUST00000140332"; exon_number "3"; gene_name "Crem"; transcript_name "Crem-025";


After encountering several of these GFF warnings:

GFF warning: merging adjacent/overlapping segments of ENSMUST00000064220 on 11 (120012520-120012843, 120012846-120012875)
GFF warning: merging adjacent/overlapping segments of ENSMUST00000106216 on 11 (120207664-120207738, 120207740-120207847)
GFF warning: merging adjacent/overlapping segments of ENSMUST00000066629 on 11 (120488995-120489234, 120489237-120489272)

my cufflinks run stopped without producing any output.

There seems to be an upper limit of warnings/errors cufflinks can tolerate before terminating the process itself.

Is this normal ? How do people avoid such errors ?

In addition, when I tried to run the same data without a GTF reference as in

cufflinks -p 16 -o ./outputdir accepted_hits.bam

This came up :

terminate called after throwing an instance of 'std::bad_alloc'
what(): St9bad_alloc

This seems to be a hardware / memory issue, any ways to sidestep this ?

Your help will be greatly appreciated.

[ayfchang]

You can solve the first problem by removing gene entries with introns larger than a certain size in the reference GTF file.

The default settings in cufflinks expect intron not larger than 300,000 bp unless you specify otherwise.

The reference GTF used here has some entries with intron size exceeding that limit.

You then re-run cufflinks -G with the newly edited GTF reference.