Frequently Asked Questions at the NMMSU Flyshop: Q: My injections were healthy and I had lots of fertile crosses, why didn't I get any transformants? A: 1. The DNA you gave us isn't what you thought it was...Please check the sequence and look more carefully at the restriction map. The vast majority of cases where we've struck out on transformants from a reasonable number of fertile crosses there is something wrong with the DNA in the tube you gave us. This can range from defects in the vector, giving us the wrong tube of DNA, or doing something else completely unexpected. 2. Your construct is BIG. The smaller the construct, the better the rate. pUAS-T constructs that only put 7 or 8 kb into the genome have a much higher transformation effeciency than a 20 kb Casper insertion. This can be on the order of 15% vs 1-3%. Last but not least: transformation efficiency is a bell shaped curve. There are unlucky souls at the disappointing end of the curve.   Q: I gave you DNA and you injected it and everything or almost everything died. Don't you people know what you're doing? A: Well, we think we do. But usually when DNA is lethal there are two prime reasons. The first is that you made lousy DNA, sorry to blame the victim, but this happens. My recommendations are to get a new Qiagen kit and FOLLOW OUR INSTRUCTIONS. There are lots of competing methods with Qiagen, and we've never tried them, so I can't say if that cheaper Qiagen imitation won't make lethal DNA. We have gotten lethal DNA made from kits from Promega and Sigma, however. The other recommendation is to go with Tranformation Service B. It costs more, but we know how to make DNA that goes into flies and if it fails, we'll do it again. The Second reason everything died, is that the DNA you want transformed encodes something EXTREMELY TOXIC. We do our injections at room temperature, but we can do them at 18 degrees if necessary, so if you are trying to make transformants of something behind a heat shock promotor that may be nasty, tell us and we'll inject a dozen embryos and make sure they're ok.   Q: How soon will I see larvae? A: Our optimal turn-around time is 7-10 days, although now that the word is out on our service, we're getting more and more constructs in the mail everyday. We're injecting 5 days a week, now and our queue is backing up. A more realistic expectation is that you'd get your larvae about 3 weeks after we get the DNA (plus or minus 3 weeks).