Abstract

As a pre-requisite to examine the possibility of using spermatozoa as a vector for the incorporation of foreign DNA into livestock, the ability of sperm from sheep, fallow deer and cattle to bind exogenous DNA was examined. The DNA was obtained from the plasmid pMK1 linearised with EcORIXhoI and end-labelled with 32P. Fresh sperm from all species bound DNA with maximum binding occurring after 1hr of incubation. Frozen sperm bound less DNA and formaldehyde treated sperm did not bind. Seminal plasma inhibited binding as did heparin and also high divalent cation concentrations. Capacitated ram sperm bound more than uncapacitated (64% vs 40%) and uterine fluids from sheep two-days post-estrus did not interfere with DNA binding to the capacitated ram sperm. Increasing the concentration of washed capacitated ram sperm from 1 x 10 6/ml to 30 x 10 6/ml and with a DNA concentration of 0.25 mg/ml decreased the number of molecules of DNA bound from 38,000 to 1,600. There was wide variation among rams in the percentage of DNA bound after 1hr incubation, ranging from 15-80% (n=7). These results suggest that this procedure may be a possible method for the incorporation of foreign DNA into livestock by intrauterine insemination.

PF, Fennessy

Proceedings of the New Zealand Society of Animal Production, Volume 50, , 361-372, 1990