I want to construct a bicistronic vector, which can co-express two different genes in Drosophila S2 cells. One way is to add IRES sequence between two genes, how to design the IRES sequence? Is it related to the genes I want to expressed? Thank you very much.
IRESs are typically used in mammalian cells. The most common one is from EMV. EMV IRES is ~500 bp long, and you add it by standard molecular biology techniques (PCR/cloning). Please, make sure that you use IRES that has been validated in insect cells. Also 2A and 2A-like sequences may be useful. Check out this paper:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3216562/
You can easily get suitable IRES containing plasmids from Addgene. I'd advise cloning it rather than attempting to PCR amplify as this can be difficult due to a very GC rich clamp at one end. IRES or IRES2? Doesn't make a lot of difference really, either way you will get a reduction in translation of the downstream cistron, typically about 70% the level of the upstream one. Personally I prefer 2A as it is very short and can even be incorporated onto the end of a primer to facilitate PCR-based cloning. The advantage of IRES is that you don't have to worry about reading frames (but do keep the ATG at the start of your downstream cistron) while 2A has to be in-frame. Expression is more stoichiometric with 2A, however there can be a low incidence of failure to cleave between the two peptides which effectively leaves you with a small amount of fusion protein. Usually that is not a problem but it's worth being aware of. Hope that helps.
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