Hi there. I have cufflinks running without errors except it produces empty transcript.gtf files. I am aliging reads with Bowtie2 and converting sam into bam and indexing with samtools. I then take the indexed file (.sorted) into cufflinks with my fasta file and gff file. The GFF and the fasta have the correct headings (there is no separate chromosomes).
Using this command
/opt/cufflinks/cufflinks -o cufflinks/sample.gff -g genome.gff -p8 -b genome.fasta -u --library-type fr-unstranded sample.sorted --verbose
I get a bunch of warnings like this
Warning: CDS ORF_02239 found without exon segments (adding default exon). Warning: CDS ORF_02240 found without exon segments (adding default exon). Warning: CDS ORF_02241 found without exon segments (adding default exon). Warning: CDS ORF_02242 found without exon segments (adding default exon).
Kept 2279 transfrags out of 2279 1: null [14:54:58] Inspecting reads and determining fragment length distribution. Inspecting bundle null:0-2253459 with 9257857 reads Processed 1 loci. Warning: Using default Gaussian distribution due to insufficient paired-end reads in open ranges. It is recommended that correct parameters (--frag-len-mean and --frag-len-std-dev) be provided. > Map Properties: > Normalized Map Mass: 9296492.50 > Raw Map Mass: 9296492.50 > Number of Multi-Reads: 0 (with 0 total hits) > Fragment Length Distribution: Truncated Gaussian (default) > Default Mean: 200 > Default Std Dev: 80 0 ReadHits still live Found 2 reference contigs Total map density: 9296492.500000 [14:58:45] Assembling transcripts and initializing abundances for multi-read correction. null:0-2253459 Processing new bundle with 9257857 alignments Processed 1 loci. [15:02:34] Loading reference annotation and sequence. Kept 0 transfrags out of 0 [15:02:34] Learning bias parameters. Processed 0 loci. [15:03:05] Re-estimating abundances with bias and multi-read correction. Processed 0 loci.
Please can i have some help im at a brick wall.