Should we take a mRNA->cDNA route, or a complete genome analysis? Please keep in mind, that the organism is not a virus, and that the method should also be cost effective.
The term "cost effective" is confusing. If you are only interested in cloning any new gene, as the organism is novel, the cheapest would be to extract RNA, do cDNA synthesis using oligo-dT (I assume this organism has polyA mRNA) and use random hexamers to amplify the mRNA. TA clone and sequence a random clone. If you are interested in cloning as many unknown genes as possible, then I would definitely go the RNAseq route. The only problem is if there is genomic data for the organism as you will need this to anchor your short sequencing runs.
It depends on the purpose. If interested in differential expression, you can use mRNA DDRT PCR or Subtractive Hybridization. You get some 50 odd clones sequenced and you are very much likely to get few genes with unknown function. Cost effective is relative term and depends on much your laboratory can afford.
Dear Matthias Schaefer and Marisol Herrera-Rivero,
Oh, thank you for telling that the question had no thought behind it! I am amazed that you are able to think! An unknown gene here means a gene whose sequence is not known. It is just obvious that one shall not blindly go and isolate something! I meant isolating a gene (sequence unknown but product isolated, tested and characterized) from an organism (novel - in the sense, a new strain/ a species). Thank you Mr. Jaime and Mr. Sudhakar for the answers!
i guess, there should be some known about the gene; might be in other species that can give clue and ou can trace back from conserved region amplification
If completly blind, then get ready for tsting all known mrna from cDNA with sequences in ncbi, in gene network so that you can find similar genes and their functions and function of that specific gene
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