RNA structure prediction-BLAST alignment to multiple sequence alignment

Question

[d.awwad19]

Hi guys,

I know this might sound like a really basic question for some, but I'm a very beginner in bioinformatics and sequence analysis..

So, I ran blastn on NCBI-BLAST on an RNA sequence to which I intend to predict the secondary structure.. The output of the BLAST contains some sequences that are labeled "plus/plus" and some that are labeled "plus/minus"..

My question is: my next step should be multiple sequence alignment.. Should I flip the "minus" subject sequences so all the sequences in the MSA input are in the same orientation, or should I run MSA on the BLAST output sequences as they are?

Thanks in advance!

[Asaf]

The BLAST output should already be flipped. If you run RNA vs other RNA then you shouldn't expect plus/minus results, these results would only appear if your database is DNA.

[d.awwad19]

It was actually the DNA exons sequence coding for the RNA molecule, but the ultimate goal is to predict the secondary structure of the RNA molecule. So as I explained in my question, some of the output sequences are in plus/minus format.. should I flip them so that all the sequences are in the same orientation before I run MSA?

[Asaf]

If the input was the coding DNA then you shouldn't expect plus/minus results, unless you found a binding site for the RNA or something like that. I would take a second look in these results to see if they're valid. If most of your results are plus/plus (way over half which is expected at random) I would exclude the plus/minus. If you get about half plus/plus and half plus/minus recheck your dataset.