Dynamic changes in microtubules and early development of reconstructed embryos after somatic cell nuclear transfer in...

In order to improve somatic cell nuclear transfer (SCNT) efficiency and to understand cellular changes in SCNT, the dynamic changes in microtubules/DNA and early development of SCNT embryos with single or multiple pronuclei were investigated, along with activation timing on efficiency of SCNT, were studied in the Cynomolgus monkey. The confocal images showed that microtubules assembled around condensed DNA at 1h after cell injection; normal or abnormal reconstructed spindle formed at 2 h after cell injection; and reconstructed spindle separated at 2 h after activation. The results of nuclear formation showed that 61.3% of the reconstructed embryos did not form pronuclei; 19.3% formed a single nucleus, and 11.9% and 7.5% formed two and more than two reconstructed pronuclei, respectively. The cleavage and 8-cell development rates of SCNT embryos with pronuclei were significantly higher than those without pronuclei, but there was no difference in development rates among NT embryos with single, two and more then two pronuclei. Activation at 2 h after cell injection yielded more embryos with pronuclei and yielded 8-cell NT embryos more reliably than did activation at 3-4 h. In conclusion, microtubules assembled around condensed DNA at 1-2 h after cell injection, and formed a spindle at 2 h after SCNT, which separated at 2 h after activation; early development was affected by activation time, but no different between single and multiple pronuclei.