Hi,
Does anyone have an idea of what factors may influence the inheritance of extrachromasomal arrays after transformation using microinjection? I would like to have an array that is inherited in about %50 of the progeny. I want to inject a PCR product as a rescue array along with a GFP marker. Any tips, tricks, or thoughts would be greatly appreciated!
There is at least an anecdotal connection between transmission frequency and DNA concentration at time of injection: a lower concentration of DNA in the injection mix is supposed to often give a lower transmission frequency. For this reason, some people bump up their total DNA concentration with extra carrier DNA, such as DNA molecular weight marker. In your case, you might try reducing your total DNA concentration.
Also remember that the array is lost in progeny because of mitotic loss in the germline, and mitotic loss is also occurring in the soma, making animals mosaic. If you want an array that’s lost in the progeny at relatively high frequency, you may also find that animals carrying the array are more highly mosaic than they would be for an array transmitted more often. Just in case that’s going to affect your plans for the transgene.
If you want 50% transmission, you could integrate your array by bombardment or microinjection/gamma irradiation. Crossing the heterozygous array line to non-array strains yields 50% transmission in outcross progeny. Plus, you don’t have to worry about mosaicism (although germ line silencing can still be problematic, but that’s also true for extrachromosomal arrays).
-Harold