I used this primer set: SC-Act-235aS20, SC-Act-878aA19, but the results are good to the genus level but not to the species level.
Did you get it from the publication of Stach et al., 2003? I am also using this primer set too with the hope of identifying to species level.
Yes I did but no success in terms of definite species level in 20 samples.
If you are using isolates the best is for you to use university primers 27F and 1525R.
Go to the Genebank for Actinobacteria and do some cross referencing. You may be able to locate one good primer. For routine bacterial identification, the universal forward and reverse primers work pretty satisfactorily. It may not be the case here. You may have to get one primer tailor made for your work.
@ Sanjay Antony-Babu. Can this primer set be used to identify actinobacteria to species level? I also need information on this.
Yes it works pretty well for species level identification for actinobacteria. This primer set (as you can see from the E.coli numbering) would give you close to 1500 bp sequence (when sequenced in both directions). This is enough to recognise most bacterial species. Remember that you cannot easily differentiate Streptomyces spp based on 16S rRNA gene sequence (whatever be your primer).
27f - 5'-AGAGTTTGATCMTGGCTCAG-3'
1525r - 5' - AAGGAGGTGWTCCARCC-3'
Original reference:
Lane DJ: 1991, 16S/23S rRNA sequencing. In: Nucleic acid techniques in bacterial systematics, ed. Stackebrandt E, Goodfellow M, pp. 115–175. Wiley, New York, NY.
You cannot identify an actinobacteria or any bacteria isolate to species-level solely rely on 16S rRNA. And in most cases, it's not even possible to distinguish close-related bacteria via 16S rRNA gene.
If you want to identify your strains, you may need to look up those articles describing new types published on Int J Sys Evo Microb
Often, bacterial species identification needs a Multilocus Sequence Alignment (MLSA), and sometimes its not enough, but needs coupling with biochemical and cultural characteristics.
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