Genetic diversity of H.pylori

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Genetic diversity of H.pylori

Postby Aschana » Feb 13 2015 1:55 am

hello, i'm studying in the genetic diversity of HP. I have used universal vacA gene to study, this gene is presented in all HP strain.
However, most of journal use vac A genotype primer (s1,s2,m1,m2) for study diversity genetic of H. pylori. But my current work use universal vac A primer that are unable to indicate vac A genotype.

So for my question,

1. Can I use this primer to study in the genetic diversity (phylogenetic tree etc.) ? :cry:
2. If yes or no, please give reasons.

thank you so much for all answer :D

Ps. my universal vac A primer covers both conserve and variable gene region and product size is 276 bps
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Re: Genetic diversity of H.pylori

Postby r.rosati » Feb 13 2015 8:11 am

Hello! Please let me understand your question: you say that most studies use allele-specific primers, but you're using a universal primer pair that spans a variable region. Is this correct? In this case, it seems logical that your work will involve Sanger sequencing of the PCR product. Or maybe restriction digestion? What's the plan for identifying the alleles after the universal PCR? This would be a legitimate question to ask to your supervisor, right? He/she should be instruct you about what you need to do, and then you're supposed to become familiar with the techniques.
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Re: Genetic diversity of H.pylori

Postby Aschana » Feb 13 2015 8:52 am

Hi, I apologize for unclear question. :(
In my work, I and my advisor want to know the genetic diversity of HP in oral cavity compared to Hp in gastric biopsy. For the first time, I used 16s rRNA of HP for amplifing product and sent them to sequence. After that, they will be taken to phylogenetic tree analysis in order to observe relationship of Hp in oral and Hp in gastric biopsy. But this primer was not specific to H. pylori. So I decide to use universal vacA primer instead of 16s rRNA due to this primer show good sensitivity and specificity.
However, when I review paper, I found the most of them use only vacA genotype primer but no one used universal vacA for study genetic diversity of Hp. So I and my advisor concern whether product of primer can be used to sequence for phylogenetic tree analysis (the objective of phylogenetic tree method, I want to know Hp in oral cavity and Hp in gastric biopsy is in the same group or not)
Finally, I hope you understand my question and thx for any suggestion :)
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Re: Genetic diversity of H.pylori

Postby mchlbrmn » Feb 13 2015 1:43 pm

Are the VacA genotype primers specific, so they only amplify one genotype? If so, wouldn't a universal primer be better for a phylogenetic tree analysis? Do I understand you?
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Re: Genetic diversity of H.pylori

Postby Aschana » Feb 13 2015 2:04 pm

I think I understood your answer. You mean my universal vacA primer be better than vacA genotype primer due to they not only amplified one genotype so they are suitable for phylogenetic tree analysis, didn't you?
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Re: Genetic diversity of H.pylori

Postby mchlbrmn » Feb 15 2015 12:10 pm

Yes, although I'm not expert in this field, and I don't know the specifics. If other people used the "specific" genotype primers for this purpose, perhaps they're not so specific, but are universal for a subset of H pylori? So maybe the set of genotype primers used together are more "universal" than the one set of universal primers? I don't know; I'm just guessing. Hopefully the literature will make this clear to you. Another guess is that they used the genotype primers instead of universal primers so they could break the diversity down into subsets of sequences, so that if, for example, one sequence is very common and comprises most of the sequences, they won't have to look at a huge number of sequences to see the rare sequences, but the other "group" primers won't "see" this sequence at all, and can find more rare sequences more quickly.
Or, does the "diversity" of H pylori only consist of four groups, s1, s2, m1, m2? Or, that's only what the other groups were looking at?
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