Quick sex determination of mouse fetuses

We designed a rapid, simple and accurate PCR method to determine sexual ide ntity of mouse fetuses collected on embryonic day 15. A multiplex PCR ampli fication was used to detect male-specific sequence (Sry) in DNA extracted f rom fetal livers through SDS denaturation followed by high salt extraction and precipitation. This extraction method resulted in sufficiently purified DNA in < 1 h and was suitable for PCR. The DNA obtained was amplified usin g a robot thermal cycler for 33 cycles. The reaction was performed in 50 mu l, using two sets of primers specific for Sry gene (chromosome Y) and IL3 gene (chromosome 11). Amplification duration was 1.5 h. The assessment of t he results was done by electrophoresis in 3% agarose run at high voltage. T he 402 bp band (Sry) obtained identifies the male fetuses and the 544 bp pr oduct (IL3) confirms the correct amplification of the template DNA. The ent ire procedure took < 4 h. The specificity of the method was confirmed by fl uorescent in situ hybridization using a specific male probe on cultured mal e and female neural stem cells. This method allowed the preparation and cul ture of pure male and female neural stem cells from fetal tissue. (C) 2000 Elsevier Science B.V. All rights reserved.