The goal of my subproject is to find transcripts upregulated upon treatment in the larvae of Spodoptera littoralis. On the current step I have an assembly, and the tblastx results (assembly against the huge database from NCBI). In order to continue with annotation I transformed blast results into gtf file (wrote a python script that also does cutoff and then write results into gtf format). The gtf file looks like this:
Slitt_C1 tblastx exon 4225 5697 667 + . gene_id "gi|827554818|ref|XM_004929801.2|"; transcript_id "gi|827554818|ref|XM_004929801.2|";
After that aligned the reads to the assembly by bowtie2, and tried to run tophat2.
~/bin/tophat2 -G bowtie_result/Slitt.gtf -o tophat_with_annotation/ -p 16 bowtie_result/Slitt ../02_trim/A_R1_P.fq ../02_trim/A_R2_P.fq
And I got the error:
[2016-07-28 11:05:14] Beginning TopHat run (v2.1.1) ----------------------------------------------- [2016-07-28 11:05:14] Checking for Bowtie Bowtie version: 126.96.36.199 [2016-07-28 11:05:15] Checking for Bowtie index files (genome).. [2016-07-28 11:05:15] Checking for reference FASTA file [2016-07-28 11:05:15] Generating SAM header for bowtie_result/Slitt [2016-07-28 11:05:15] Reading known junctions from GTF file [2016-07-28 11:05:20] Preparing reads left reads: min. length=59, max. length=99, 4805446 kept reads (1341 discarded) right reads: min. length=59, max. length=99, 4795101 kept reads (11686 discarded) [2016-07-28 11:08:38] Building transcriptome data files transcriptome_index/Slitt [2016-07-28 11:08:42] Building Bowtie index from Slitt.fa [FAILED] Error: Couldn't build bowtie index with err = 1
After looking into the log file, the last thing tophat was trying to do was to run bowtie2-build on the Slitt.fa from the temp folder.
I already checked the names in the assembly and annotation, it is all the same, so mistake is coming from something else. I would appreciate any tips how to get blast results into the expression level file (of course I can run a script for assigning the regions of scaffolds to the specific annotations, but it will take a lot of time).