I would like to study the microbial biodiversity of a hypersaline water by PCR-DGGE followed by sequencing. How to choose the suitable primers for both archaea an bacteria.
Hi Beddal,
Unless there is a reason you are using DGGE (e.g. save money while you search for an interesting site), I would just go straight to the 16S rRNA sequencing. Also, unless you have a reason to separate bacteria from archaea, I would use universal primers. Given your site, I would recommend that you use primers well tested from the Earth microbiome project: http://press.igsb.anl.gov/earthmicrobiome/protocols-and-standards/16s/
JGI:
http://jgi.doe.gov/wp-content/uploads/2013/05/iTagger-methods.pdf
If you want closer to full-length 16S sequences, then you might have to separate bacteria and archaea, but the primer research is out there in the literature.
You can always reach out to your sequencing center of choice for more specific questions about the primers you choose.
I agree with Dr. Eric Daniel Becraft's answer. I think, 16S rRNA gene sequencing is the best option to study microbial diversity. Please see this list of primers for halophilic bacterial and archaeal diversity analysis from hypersaline environments.
I partly agree with Eric .His point of view is based on the affordability, which is an important issue in many such experiments. However, because of rampant use of this technique of 16SrRNA sequencing to study the phylogeny, I feel that it is being used in many wrong homology studies. The result is that the outcome in many of these cases are erroneus. Now many scientist do not accept this phylogenetic analysis and are asking for entire genome sequence-homology studies to get better and reliable results.
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