getting reference for indexing the transcriptom needed for tophat2

Question

[zizigolu]

Sorry dudes

Newly I recognized that I should index the transcriptome, from where I can get the gtf file for yeast please?

Thank you

Answer 1

[zizigolu]

http://www.ensembl.org/info/data/ftp/index.html

[Ashutosh Pandey]

Its always good to do things from scratch (starting with the raw file) because that way you learn lot more. However if you don't feel like doing it, you can download pre-processed genome and transcriptome index, and annotation files from iGenomes. They have one for yeast too. Check here: https://support.illumina.com/sequencing/sequencing_software/igenome.html

[zizigolu]

Thank you dear ashutosh

[zizigolu]

sorry,

in https://support.illumina.com/sequencing/sequencing_software/igenome.html there are

• R64-1-1
• EF4
• EF3
• EF2

then which I should download maybe it is match and compatible with my reference genome???? I am really confused...

in http://www.ensembl.org/info/data/ftp/index.html there many chromosore in DNA (fasta)...then I donno finally how I can have the indexed transcriptom matched with my reference

please help me

p

[Ashutosh Pandey]

They also provide the reference genome (which should be the most latest one) and its Bowtie2, BWA and Tophat2 Index. So its a pretty much one stop shop for NGS analysis. R64-1-1 is the most latest reference genome, so download the whole folder. It may take some time to download it so have patience but it will save your time building index for Tophat2. It also provides the genes.gtf file that can directly go to Tophat along with the Tophat2 index.

[zizigolu]

thanks a lot Ashutosh,

this is my result

Reads:
          Input     :  16286258
           Mapped   :  15989053 (98.2% of input)
            of these:   7149891 (44.7%) have multiple alignments (51923 have >20)
98.2% overall read mapping rate.

what can I do please to reduce multiple alignment rate...my adviser asked me to try to do nearly unique alignment