I’ve been recently using the BLAST+ blastn tool to search my de novo assembled contigs for sequences of interest. I’ve done this successfully with both gene and primer sequences.
However, I queried these same contigs (which had been made into a database with “NCBI BLAST+ makeblastdb”) with primer sequences I expected to find in a subset of the contigs and the blastn output was blank. When I pull out an individual contig that I suspected to have the sequence, turn it into a database, and run the blastn, the result shows a perfect match.
It seems like the issue may lay somewhere with BLASTing multiple sequences at once with these particular primers (since I’ve done this same workflow with identical settings successfully with other sequences including the gene that the primers were designed for).
Anyone run into this same issue/have any advice?
I apologize if any important details are missing, please let me know what else would be needed.