18S sequence not matching 18S sequence in assembly
0
0
Entering edit mode
16 months ago

I have a genome assembly that I used to make a blast database. I then took the 18S sequence and blasted against the database. I got results showing contigs that had the 18S matching 100% identity, with 0 gaps and 0 mismatches. Once I extracted the contig and aligned the 18S sequence in Clustal- I see several gaps and mismatches. How can that be?

BLAST database 18S Genome assembly • 389 views
ADD COMMENT
0
Entering edit mode

did you extract the whole contig or only the region indicated by the blast result? in the latter, what you describe should not happen, in the former it could as clustal is a global aligner and will thus try to align the two sequences completely resulting a gapped alignments (and likely not very accurate).

ADD REPLY
0
Entering edit mode

Initially I was extracting the whole contig. I just tried it now extracting 2500 bases from the contig and aligning using clustal omega and still not seeing the 100% seq identity as reported in the blastdb results.

ADD REPLY
0
Entering edit mode

I used the contig to make a blast database and then blasted using the 18S sequence and now the sequence identity is only 92% which matches up with the results I was seeing when I aligned the 18S with the whole contig or the part of the contig that aligned to the 18S.

ADD REPLY
0
Entering edit mode

ok, so that already adds up

in your original blast was the hit on single match on a single contig? It could also be that blast did not align the complete 18S sequence and only reported the 100% match region?

Can you perhaps post an extract of that orginal blast result?

ADD REPLY
1
Entering edit mode

I figured it out. I extracted the contig from a contig.fasta file but then when I went to make the database of the full assembly I was using the assembly.fasta file. When I use the same assembly file to extract the contig and to make the database of the assembly they match 100%. I didn't realize the contigs in these files would be different.

ADD REPLY
0
Entering edit mode

This is the resulting output of taking the 18S and aligning it against my assembly db using blastn https://ibb.co/r3Ynqk9

This is the resulting output of taking the 18S and aligning it against contig 672 (which I made into a blastdb) which previously had an identity of 100% https://ibb.co/gmL7SS3 The headers for the output are: # Fields: query id, subject ids, query acc.ver, subject acc.ver, % identity, alignment length, mismatches, gap opens, q. start, q. end, s. start, s. end, evalue, bit score

ADD REPLY

Login before adding your answer.

Traffic: 2053 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6