I want to do microbial community identification by culture-based method and molecular based techniques.
I will start with bacterial Identification then fungal identification, so what are the best biochemical characterizations i should do for bacteria?
There are some biochemical characterizations commonly used for bacteria , such as colony morphology, gram test, carbohydrate fermentation determination, you can follow Bergey's manual of determination bacteriology.
For identification, some molecular biological identification methods are needed, e.g. DNA extracts, PCR, and 16S rDNA .
Good luck!
J.G. Holt, N.R. Krieg, P.H.A. Sneath, J.T. Staley, S.T. Williams Bergy's Manual of Determinative Bacteriology (ninth ed), Williams and Wilkins Pub., MD, USA (1994)
Dear Eman,
Yuan gives you nice suggestion.
For the identification of isolated bacteria from the soil you have to go for morphological, biochemical, physiological and phylogenetic (16S rRNA or rDNA) characterization.
If you want to do the total microbial identification of soil, follow soil DNA extraction and then PCR-DGGE, pyrosequencing etc. for the identification of different microorganisms in soil.
For fungal identification: morphological, microscopic characterization and molecular (ITS characterization) can be performed.
Kunal
Dear Kunal,
Thank you very much for your reply.
I want to know more details about the biochemical characterization of bacteria and universal primers for PCR and sequencing.
Many Thanks.
Eman
Hey
One of the best ways for biochemical analysis is using the API identification method:
http://www.biomerieux-usa.com/servlet/srt/bio/usa/dynPage?doc=USA_PRD_LST_G_PRD_USA_5
In this strips you have several nutrients and products and consequent inoculation of your strain will lead to a result that you'll be able to compare and obtain a probabilistic identification of your strain.
The other step would be sequencing of the 16S and 18S ribossomal gene (for bacteria and fungi respectively) and analysis with gene-databases.
Dear Eman,
Biochemical characterization can be done manually (procedure from the literature published such as in International J of Systematic and Evolutionary Microbiology, and many more) or by using BD Phoenix™ Automated Microbiology System (link attached) or similar techniques.
For 16S rRNA or rDNA you can use several reported universal primers(http://en.wikipedia.org/wiki/16S_ribosomal_RNA) or form the literature published.
Hope this will help you.
Kunal
https://www.bd.com/ds/productCenter/IS-Phoenix.asp
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