One of my 16S rRNA gene sequencing resulted in 93% similarity to NCBI database., suggesting it could be a novel or new species., which im not sure yet.
For this case, what should i further do ?
Whole genome sequencing? or DNA-DNA Hybridization? or anyother analysis?
Kindly recommend further analysis and also, where exactly can it b done?
Thanks in Advance
Lokesh
At least 1400 bps is enough then you can go for further steps including drawing the phylogenetic trees within the family of Streptomycetaceae. If the similarity value by Ez-taxon is below 97% then you do not need to do DDH. You should do morphological and chemotaxonomical characterisation to confirm the novelty of your isolate.
The first step would be determining the the full length of 16S rRNA gene. then check the similarity value in Ez-taxon server. based on the results of 16S then you can go further with more steps.
I have a sequence of 1401bp length. How much bp does it roughly take for a complete 16S rRNA gene? I am not getting a definite answer on web. .
All of the 16S rRNA gene in GenBank sequence for Streptomyces, are mentioned as partial.. How long is a complete gene?
At least 1400 bps is enough then you can go for further steps including drawing the phylogenetic trees within the family of Streptomycetaceae. If the similarity value by Ez-taxon is below 97% then you do not need to do DDH. You should do morphological and chemotaxonomical characterisation to confirm the novelty of your isolate.
You can please refer to the links below to obtain certain valuable informations:-
1. http://www.ncbi.nlm.nih.gov/pubmed/15989219
2. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3786795/
3. http://www.sciencedirect.com/science/article/pii/S0944501305000844
Please check this reference. 16S is not always optimal.
http://www.ncbi.nlm.nih.gov/pubmed/24153003
Dear Lokesh,
have you carried out 16S rDNA sequence analysis at EzTaxon site. any seq. greater than 1400 nucleotides is OK. so, your seq. length is alright. look for its quality as well, if that is alos oK then what is the closest match and what is the seq. similarity. Remember, if your seq. identity is less than 98.8% (earlier it was 97%), very likely your strain is a novel sp. of Streptomyces.
To prove that you have to carry out detail comparative phenotypic characterization with close phylogenetic relatives. this includes morphology, biochemical tests, growth on ISP medium (imp for Streptomyctes), FAME, Menaquinone, G+C, Cell wall type analysis, glycoyl type etc. To come to conclusion. All these tests are highly specialised and you need a specialist to consult.
You can contact Dr. Mayil Raj, He is a Scientist at MTCC, IMTECH, Chandigarh, you can go to website of IMTECH, Chandigarh and get his email & ph no. and can contact him.
He is a specialist in Actinobacteria and right now the best person in India.
Hope it will give direction if your strain is novel.
Remember, if you have novel function (some antimicrobial) go for patent. In that case you have deposit your culture in a IDA (international depository authority) under safe deposite, In India, there arre two MTCC, IMTECH Chandigarh and MCC, NCCS, Pune.
Best Wishes
Pradipta
Please add to my earlier answer that if your sequence identity is below 98.6%, DNA-DNA hybridization experiment and Delta Tm determination may not be required if there is sufficient differences in comparative chemotaxonomic and phenotypic characters with its close phylogenetic neighbours.
Please add
https://www.researchgate.net/post/When_I_can_say_that_a_novel_bacterial_strain_has_been_isolated2
Dear Lokesh,
your strain can well be a new genus. a thorough analysis is required. Streptomyces is a tight cluster represented by largest asembledge of known spp (nearly 500 or more). You do not have to carry out DNA-DNA hybridization. what you have to carry out is detail polyphasic taxonomic characterization using nearest phylogenetic neighbours. You have to do a detail chemotaxonomic analysis (Cell wall type, menaquinone, glycoyl type in addition to FAME), if you do not have expertise to do these contact Dr. Mayilraj of MTCC, IMTECH, Chandigarh, India. He is a Scientist of Actinobacteria section and is right now the best person in India, to solve your problem.
Pradipta
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