Direct comparison between ICSI-mediated DNA transfer and pronuclear DNA microinjection for producing transgenic rats.

Production efficiency of transgenic rats was compared directly between the routine pronuclear microinjection of exogenous DNA solution (PNMI-Tg method) and the ooplasmic injection of sperm cells exposed to exogenous DNA solution (ICSI-Tg method) using six DNA constructs. The overall production efficiency per treated oocyte in the ICSI-Tg method (mean 1.1%, range 0.2 to 3.1%) was similar to that in the PNMI-Tg method (mean 1.1%, range 0 to 2.4%). An advantage of the ICSI-Tg method in the production of transgenic rats is noted in cases in which a low yield of pronuclear zygotes is an inevitable fate of the rat strain.

Contribution of spermatozoal centrosomes to the microtubule-organizing centre in Antarctic minke whale ( Balaenoptera bonaerensis ).

Using an interspecies microinsemination assay with bovine oocytes, it was examined whether centrosomes of Antarctic minke whale spermatozoa function as the microtubule-organizing centre (MTOC). Bull and rat spermatozoa were used as positive and negative controls, respectively. Vitrified-warmed bovine mature oocytes were subjected to immunostaining against alpha-tubulin 4-6 h after intracytoplasmic injection (ICSI) of 5 mM dithiothreitol-treated spermatozoa. Aster formation occurred from whale spermatozoa (33%) and bull spermatozoa (33%), but very little from rat spermatozoa (3%). Activation treatment for the microinseminated oocytes with 7% ethanol + 2 mM 6-dimethylaminopurine resulted in a similar proportion of oocytes forming a whale sperm aster (35% vs 27% in the non-treated group; 4 h after ICSI) but a significantly larger aster (ratio of aster diameter to oocyte diameter, 0.57 vs 0.30 in the non-treated group). These results indicate that the centrosome introduced into bovine oocytes by whale spermatozoa contributes to the MTOC and that assembly of the microtubule network is promoted by oocyte activation.