How to add GFP in STAR reference?

Question

[bioinfo]

Hello,

I wanted to ask how to add GFP to a STAR reference. I have added the GFP fasta sequence to the human fasta file. Then, I want to add the GFP to the gtf.

I was going to add the below to the end of the .gtf file. Do I need to know anything about the strand? Is there a chance I would need to use the "-" for reverse strand?

GFP     unknown exon    1       580     .       +       .       gene_id "GFP"; transcript_id "GFP"; gene_name "GFP"; gene_biotype "protein_coding";

Thank you

[GenoMax]

It may be best to include a gene entry in addition so the file matches GTF spec.

GFP     unknown gene    1       580     .       +       .       gene_id "GFP"; gene_name "GFP";
GFP     unknown exon    1       580     .       +       .       gene_id "GFP"; transcript_id "GFP"; gene_name "GFP"; gene_biotype "protein_coding";

Make sure your chromosome name for GFP in genome file matches what is in the GTF file.

[lieven.sterck]

Not a strict requirement as far as I remember, I think it will even except a . (dot) for the strand (== unstranded feature)

I would assume the GFP fasta is in sense format so + should be OK , that it is anti-sense is not impossible but not likely

[bioinfo]

Thank you. If I put the wrong stand will I just not get any counts for GFP?

[lieven.sterck]

Not even I think, it will just report that reads were counted in the opposite strand, but do check 'cus if you do really stringent counting/read-assigning they might indeed not be counted in the end result.