I am trying to get full sequence of 16S rRNA gene from actinomycete isolates. I have tried 8F and 1492R set prime but I have got 1384 bps and is still lower than about 1490 bps. I was wondering if anyone could tell me what is the best approach to reach to the full sequence of the gene?
I have never had good luck with the 8F x 1492R combination, but the 1391R reversed primer worked much better for me. It's not going to give you the full length gene, but about 90% - which is often good enough for detailed phylogenetic analysis.
The different species may contain various length of 16S rRNA gene. don't worry about your 16S length, if you have already gotten a good amplification. 8F and 1492r is the longest universary primer pair for 16S amplification.
For me primers pAF (5'- AGA GTT TGA TCC TGG CTC AG - 3') and pc5B (5'-TAC CTT GTT ACG ACT T-3') worked very well.
Dear Rodrigo tank you for your answer.
May I ask you how many sequences did you get. what bacteria you were looking for?
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