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Hai, anybody please help me to solve my problem.
My problem is
Can I use a universal primer set ( i.e., containing adaptor sequence, specific bacterial barcode sequence) either directly to amplify bacteria present at particular human habitats in normal PCR or to pyrosequence amplified PCR products ( with conventional universal primers)? . I found that primerset in the publication 'Error-correcting barcoded primers allow hundreds of samples to be pyrosequenced in multiplex', I think, there is no need to design primers again for my study. I want to identify total bacteria found in particular human habitats. Please help me.
Dear D.Esther Lebonah,
I am sorry to see that you are still struggling with that question.
Can you answer the following questions:
1. Can you make an estimate to "what kind" of bacteria you will encounter in "particular human habitats" ?
2. Did you find any publications successfully using the primers (the part that is specific to the 16S rDNA) in that group of bacteria ?
Thank you sir for taking time for me again. Sir, I've followed all of your suggestions but still I'm really struggling because I'm doing this for the first time in my department and I have no seniors to help me in this way. I want to identify all the bacteria found in human milk and in infant feces by using Barcoded pyrosequencing. But I've failed to decide universal primers for my study. I found a journal on bacteria and they provided primer set also, but I don't know how to use them directly. Can I use that primer set (which they represented in supplementary data) either directly to amplify bacteria present in human milk and infant faeces in normal PCR Or to pyrosequence amplified PCR products ( with conventional universal primers)?. I think, there is no need to design primers again for my study. I found this attached publication. It is my problem.
D. E. Lebonah
It seems like you are as alone on this as myself - I am in plants not bacteria. The question if you can use a primer for the bacterial communities you are interested in, can be answered by finding evidence of success in the literature using that particular primer sequence (the 16S rDNA specific). If you can use the primers for the approach you are planning can be answered similarly by comparing studies, which use them in a similar context. It should also be possible to combine a different specific part of the primer with the adapter and the error correcting tag. It might be a good idea to find a better place to ask this questions as it seems that nobody from the bacterial community is actually reading this forum. You could try researchgate.
O.K sir, I'll surely try researchgate today. Whenever you get a little bit more knowledge on this topic in future, please suggest me. I wish you all the best for your bright future and I hope you'll surely get success in your work.
The same to you!