Hello,

I am having low unique mapped read. I am a beginner and I might have done any mistake.

So first I downloaded a reference genome file and annotation file from ensemble.

https://drive.google.com/drive/folders/1goBTntkXcH3sKGAkrK-piSVII5FdRcUa

then I did genome indexing with the following code

STAR --runThreadN 64 --runMode genomeGenerate --genomeDir STAR_folder --genomeFastaFiles Mus_musculus.GRCm39.dna.toplevel.fa --sjdbGTFfile Mus_musculus.GRCm39.103.gtf

Next the alignment was done for this sample SRR1552444.fastq.gz

With this code:

STAR --genomeDir STAR_folder/ --readFilesIn SRR1552444.fastq.gz --readFilesCommand zcat --outFileNamePrefix STAR_Align/444_ --outSAMtype BAM SortedByCoordinate --runThreadN 16 --sjdbGTFfile Mus_musculus.GRCm39.103.gtf

The problem was that the unique mapped reads were only 15% and from literature it is a least 75 %

Do you know where the mistake might be?

Best Regards,