When analysis bacterial DNA sequences the consensus doesnt match the reference sequence, when blasting them. Many bad psectra are viewed in the analysis software. Please what to do in this case?
Is this with NGS or Sanger sequencing. If Sanger is the signal strength strong and are the badly defined sequence regions at the start and finish of the sequence trimmed off before analysis?
I am assuming you are working with Sanger sequencing and you are analyzing the quality of your lectures in a software such as Chromas:
http://technelysium.com.au/wp/chromas/
If that is the case you could trim the first or the last nucleotide positions (where usually low quality is) until you remain with a full high quality sequence. Otherwise, if there are a lot of low quality lectures across your sequence you must repeat PCR and sequencing.
Luis Rodrigo Arce-Valdés thanks for your answer actually I'm using BioEdit as a software.
Manel Merradi
You are wolcome! BioEdit should also give you the option to trim your sequence to avoid low quality lectures. Good luck!
Hi, I've also encountered a similar issue. In our lab, the problem may be in the form of:
1. Mixed/low quality base at 30-50 nt of the start and the end of sequencing result. This is relatively common, you may trim those bad peaks before performing blastn.
2. Mixed base at the entire sequence. This may be caused by the presence of multiple PCR products. To avoid this, you may use sequencing service using gel cut.
3. Be aware of homopolymer sequence above 10 nucleotides, which may cause polymerase slippage. To avoid low quality sequencing result, you may proceed your sequencing from reverse or even both directions.
I would also suggest to refer to the troubleshooting guidelines if the problem still persist
You could try DNADragon available from:
https://www.sequentix.de/software_dnadragon.php
or:
https://www.dna-dragon.com/
Either use automatic end trimming or manually trim the ends ...
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