Hi All,
We have been using the PolF and PolR primers (Poly et al., 2001) to identify nitrogen fixers but unfortunately only ~ 10% of the isolated bacteria are giving the correct 360 bp band...........We are trying to optimize the conditions of PCR but we have only met with limited success this far.
Can anyone suggest a better pair of universal primers that target the nifH gene ?......the corresponding article would be helpful too.
In this paper - http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0042149 -several combinations are given but many have irregular nucleotides such as 'I'..........
Any advice on this matter would be very much appreciated.
Thank you.
Best,
Dilantha
Jonathan Zehr did a lot of work on inviromental nifH. Look at his publications/webpage:
http://www.jzehrlab.com
Yoram
Not all the Nirtrifix bacteria share same sequence of Nitrogenase, Several PCRs should perform to identify all Nirtrifix bacteria's and If you suppose to identify at once use multiplex PCR.
Need any info go through these articles.
http://aem.asm.org/content/67/12/5791.full
http://www.omicsonline.org/development-of-molecular-identification-of-nitrifying-bacteria-2155-6199.1000211.pdf
http://www.ncbi.nlm.nih.gov/pubmed/11835531
Thank you for your advice. I will see what can be done.
Best,
Dilantha
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