Microinjection vs. ballistic transformation

The Hyman Lab publication regarding the TransgeneOME project, led me to get some constructs from them. http://www.ncbi.nlm.nih.gov/pubmed/22901814

They suggest bombardment for transformation. I was wondering if I could not use microinjection? In an email, I got the reply that “microinjection will only give me arrays, whereas bombardment is going to give me stable strains”.
Can someone explain to me why? I don’t understand.

I don’t have experience with neither technique, but for the microinjection I am at least set up instrument wise… :wink:

The two big differences between microinjection and microparticle bombardment - the differences in terms of outcome, not in terms of the procedures involved - are copy number within the transgene (which will be vastly higher for an injected than a bombarded transgene) and that injected transgenes are extrachromosomal, while a large proportion (something like half?) of transgenes obtained by microparticle bombardment are integrated into a chromosome. It is possible to integrate a transgene generated by injection into the genome, by causing double-strand breaks and looking for F2 clones unable to lose the transgene, but it is an additional step that takes a fair amount of time and plates.

Hi,

Hillel has summarised nicely the main differences…just to add,

Judith Austin’s group reported that (depending upon the vector) between 9-35% of transformants from biolistic transformation were low-copy, integrated.

see: http://www.genetics.org/content/157/3/1217.long

What seems to make biolistic transformation more effective in this sense, is the likelihood that your construct gets into the germline nucleii.

I have bought a biolistic transformation system for my lab and am currently setting it up. Cost: the gun (from Ralf Schnabel not BIORAD) cost about €5K, gas, vacuum pump etc, add another €1K. Ralf’s group do a kind of ‘getting to know your helium gun’ practical session, but unless you live here in Germany or fancy a holiday that’s not much use. However, the forum must have lots of members who have used biolistic transformation.

Regards

Steve

Hi,

Thanks to both of you, and sorry for my late reply. Alright, I understand the advantages of bombardment now. I happen to live in Germany, but will ask around for people doing gene bombardment first. Just in case - has this “getting to know your helium gun” session taking place already?

Cheers,
Mindo

Hi,

no, not yet…I need to pick up the helium gun from northern Germany (I hope the secret service are not monitoring these communications… :-).

I should have it in my lab by mid-May. Depending upon where you are, it might be easiest for you to arrange to visit Ralf’s facility, or, as you say, to talk to others doing the same .

Steve