DNA-FINGERPRINTING TO DETERMINE PATERNITY IN LABORATORY RAT SPERM COMPETITION EXPERIMENTS

Prior to this study a significant amount of research had been undertak en in the field of sperm competition in mammals. However males of diff erent strains have been required in each of these studies to enable pa ternity assignment through gene expression, which has consequently res ulted in problems with differential fertilising capacity being encount ered. In this study paternity assignment of progeny from sperm competi tion experiments with Sprague Dawley rats was achieved by multilocus D NA fingerprinting using band locus matching of individual specific ban ding patterns between progeny and parents. Trials with 4 restriction e nzymes and 5 digoxygenin labelled probes (4 oligonucleotide and 1 clon ed) achieved the highest levels of DNA fingerprint heterozygosity usin g AluI(CAC)(5) and HinfI(CAC)(5) combinations; however, paternity coul d not be determined in all offspring, due to a higher than expected de gree of inbreeding within the rat population used in this study. This was demonstrated in subsequent comparisons of genetic diversity of thr ee laboratory rat breeding populations from two different animal breed ing facilities. Data from the rat mating study showed that, under cond itions of direct sperm competition, second males given access to a mat ed oestrus female either 0.5 or 6.0 h after the first mating consisten tly required less time than the first to ejaculate: 7.6 min vs. 19.5 m in (0.5 h delay); 7.8 min vs. 19.5 min (6.0 h delay). A second male si ring advantage was identified using DNA fingerprinting in both delay g roups for those offspring on which paternity could be determined: 0.5 h delay, 1st = 39%, 2nd = 61%; 6 h delay, 1st = 34%, 2nd = 66%.