Injecting a strain already carrying an array

Model Organism: Worms Methods & Reagents
1

I was wondering if anyone has tried injecting a strain already carrying an array with additional DNA (i.e. to make a second array).

2

I haven’t done this, precisely, but some relevant points:

  1. I’ve done a fair number of injections into animals with an integrated high-copy-number transgene, successfully.

  2. Having said that, some attempts I made to inject a transgene into a strain with a closely related transgene (starting with an integrated gene-1::gfp transgene and injecting a gene-1::dsRed transgene) gave lines (by co-injection marker) and retained expression from the integrated gfp reporter but did not give fluorescence from the new transgene (which did express when injected into the wild type). Discussing this with people, I heard this and related phenomena had been encountered before; mention was made of cosuppression in plants, and of the work of Alla Grishok, as possible mechanisms. This was some time ago, and I seem to recall there have been other reports since then of related phenomena with some mechanistic explanations.

3

I could see running into problems with more than one extrachromosomal array. There are reports that arrays are treated like an extra X chromosome during crosses (I can’t seem to find the link but it might be tucked away on WormMethods somewhere). This is why it can be very easy to cross the transgene into males, but very difficult to get that transgene back into a hermaphrodite (something I’ve certainly confirmed). I imagine these difficulties apply when managing two different transgenes.

4

We’ve done it on quite a few occasions with no particular problems.

5

Thanks for the replies. I have been considering making some array lines using the MosSCI plasmid (ie: pCFJ350), performing some screens on the array line, and then injecting the Peft-3::transposase later as needed for a subset of array lines to achieve integration. I am still not sure if this would actually work, but I will update this post if I decide to try this out.