Hi all,
I’m experiencing a difficulty in getting a complete bacterial genome from PacBio sequencing data using the SMRT Analysis protocols.
My usual workflow is as follows:
De novo assembly by HGAP2 ->Circularization by circlator or minimus -> Polishing by RS_Resequencing.1 protocol until no variants are detected.
This workflow has worked nicely. Usually, repeating RS_Resequencing several times resulted in 100.0% of Consensus Concordance and no variants.
But, for a new bacterial genome we sequenced recently, the result of RS_Resequencing is different from usual. During repeating the RS_Resequencing 8 times, variants have been detected continuously at 10-20 positions and Consensus Concordance never reached 100.0%. At some positions, 1-bp insertion occurred repeatedly per each resequencing.
In my opinion, it seems that more repetition of RS_Resequencing would not be helpful.
What should I do to solve this problem?
Does anyone have a similar experience?
Thanks in advance!
I’m experiencing a difficulty in getting a complete bacterial genome from PacBio sequencing data using the SMRT Analysis protocols.
My usual workflow is as follows:
De novo assembly by HGAP2 ->Circularization by circlator or minimus -> Polishing by RS_Resequencing.1 protocol until no variants are detected.
This workflow has worked nicely. Usually, repeating RS_Resequencing several times resulted in 100.0% of Consensus Concordance and no variants.
But, for a new bacterial genome we sequenced recently, the result of RS_Resequencing is different from usual. During repeating the RS_Resequencing 8 times, variants have been detected continuously at 10-20 positions and Consensus Concordance never reached 100.0%. At some positions, 1-bp insertion occurred repeatedly per each resequencing.
In my opinion, it seems that more repetition of RS_Resequencing would not be helpful.
What should I do to solve this problem?
Does anyone have a similar experience?
Thanks in advance!
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