DNA preparation method can influence outcome of transgenic animal experiments

In our continuing quest to improve the efficiency of producing transgenic a nimals, we have compared the influence of two transgene purification techni ques on the efficiency of creating transgenic sheep and mice. Three hundred eighty-seven sheep zygotes and 2,737 mouse zygotes were microinjected with one of four transgenes. Transgenes were isolated from plasmid sequences ei ther by agarose gel electrophoresis followed by gel extraction or by a sing le step sodium chloride gradient fractionation technique. Four transgenic s heep and 61 transgenic mice were produced. Both sheep and mice embryos resp onded similarly to transgene preparation methods. Overall, pregnancy rate w as higher for recipients that received embryos injected with NaCl purified DNA (mean +/- SEM: 64 +/- 7% vs. 38 +/- 7%). Furthermore, offspring per zyg ote transferred (NaCl, 22 +/- 3% vs. Gel, 12 +/- 3%) and transgenics born p er zygote transferred (NaCl, 3.9 +/- 0.6% vs. Gel, 1.5 +/- 0.6%) were highe r when the NaCl purified DNA was used. However, the proportion of offspring born that were identified as transgenic did not differ between transgene p urification methods. Transgenes responded differently to methods of prepara tion. One of the four genes yielded a significantly higher proportion of tr ansgenics when the transgene was prepared by NaCl purification. These data suggest that on average the NaCl gradient purification technique results in a higher embryo survival rate to term for both sheep and mice, but the tec hnique has no influence on rate of transgene integration.