Hi everyone,

I would like to add the sequence of a plasmid, which is integrated in a cell line that I sequenced, with more than one relevant gene to a fasta as well as gtf file and I'm not sure how to do this correctly.

If I would only be interested in one transcript of let's say GFP, as described in the example under this link (https://groups.google.com/forum/#!msg/rna-star/FGQRotrCB1Q/oQ2annphCQAJ), you would add something like this:

fasta:

">eGFP eGFP sequence"

gtf:

"eGFP AddedGenes exon 1 720 . + 0 gene_id "eGFP"; transcript_id "eGFP";"

But how would this look if I would for example like to annotate two genes from my plasmid, e.g. gene of interest and selection marker, as well as UTRs. Would I handle the whole plasmid sequence as the "gene" in the gtf and the genes I'm interested in as transcripts or would I handle the genes (selection marker and GOI) separately? But then I'm not sure what to add to the fasta files. Would I then just add the sequence of my genes + UTRs to the fasta? But then again I'm not sure what the coordinates of the respective elements would be. Maybe someone could help me by a schematic on how this would be built up correctly.

Thanks a lot!