| dc.description.abstract |
Efficient selection of preimplantation transgenic embryos by an improvedmethod after pronuclear injection of exogenous DNA is described. The method isbased on subjecting DNA extracted from the embryos to restriction enzymes aswell as the polymerase chain reaction (PCR). The incorporated procedureincluded recovery of the digested DNA with glassmilk before PCR, whichmarkedly enhanced the rate of accurate detection of transgenic embyos. Whenexogenous DNA sequences in the mouse embryos were not integrated into thegenome they were digested with both Dpn I andBal31, and subsequent PCR analysis generated DNAfragments of the injected DNA sequence in only 1 ·5% of casesexamined. However, DNA extracted from mouse embryos containing the transgenesequences integrated into the genome evaded digestion by both enzymes andyielded transgene-specific PCR products in 68· 6% of the embryostested. When bovine embryos were used, sequences of the endogenous haemoglobingene used as a control genomic DNA sequence were protected from enzymedigestion (PCR products in 70· 5% of the embryos examined); bycontrast, the non-integrated injected sequences were almost completlyeliminated by the same treatment (PCR products in 1· 4% of theembryos examined). It is suggested that this method might be useful for theselection of transgenic embryos before embryo transfer, thereby reducing thenumber of recipient females required. |
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