Effect of demecolcine and nocodazole on the efficiency of chemically assisted removal of chromosomes and the developmental potential of nuclear transferred porcine oocytes |
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Authors: | Kawakami Masahiro Tani Tetsuya Yabuuchi Akiko Kobayashi Tatsuya Murakami Hiroshi Fujimura Tatsuya Kato Yoko Tsunoda Yukio |
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Institution: | Laboratory of Animal Reproduction, College of Agriculture, Kinki University, 3327-204, Nakamachi, Nara 631-8505, Japan. |
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Abstract: | Brief treatment of metaphase II (MII) stage porcine oocytes with 0.4 microg/mL demecolcine in the presence of 0.05 M sucrose produces a membrane protrusion that contains a condensed chromosome mass. The present study examined the optimal conditions for demecolcine and nocodazole treatment in chemically assisted removal of chromosomes. When matured oocytes were treated with 0.1-0.4 microg/mL demecolcine for 60 min or with 0.4 microg/mL demecolcine for 30 min or 3 microg/mL nocodazole for 30 or 60 min, more than 70% of oocytes had a membrane protrusion containing condensed chromosomes were located. There was no difference in the in vitro developmental potential of enucleated oocytes assisted by 0.1 and 0.4 microg/mL demecolcine or 3 microg/mL nocodazole that received porcine somatic cells. After transfer to 10 recipients, however, two of six recipients that received demecolcine-treated enucleated eggs produced four healthy cloned piglets, but none of the four recipients of nocodazole-treated enucleated eggs produced piglets. Further studies are required to increase the successful development to term because the proportion of live piglets was low (4/2, 672, 0.15%). |
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