Cytogenetic Analysis of Diploidy in Cloned Bovine Embryos Using an Improved Air-Dry Karyotyping Method

Document Type

Article

Journal/Book Title/Conference

Theriogenology

Volume

63

Publication Date

2005

First Page

2434

Last Page

2444

Abstract

Of the few published studies on the cytogenetic analyses of bovinenuclear transferred (NT) embryos, results differ between air-dry and fluorescent in situ hybridization (FISH) procedures. A modified air-dry procedure is reported in this study that provides more metaphaseplates for analysis. Day 5 and Day 7 bovine NT embryos were cultured in colcemid-containing CR1aa for 10–12 or 16–18 h, then treated in hypotonic sodium citrate for 3–5 min. The standard procedure of 5 h in colcemid and 15–20 min in hypotonic solution was the control. A much higher (P < 0.01) percent of mitotic nuclei was observed in the experimental groups. The 33 and 41% mitotic nuclei were obtained from 10 to 12 h and 16 to 18 h-colcemid-treated Day 5 embryos, respectively, which was higher (P < 0.001) than the control (15%). The mitotic nuclei in Day 7 NT embryos were 24% in 10–12 h- and 28% in 16–18 h-colcemid-treated groups, which also was higher (P < 0.05) than the control (10%). The majority of analyzable embryos were diploid. Analyses of mixoploid embryos showed on average that 70% of the cells were diploid. Day 5 mixoploid embryos contained numerically higher polyploid cells than Day 7 embryos, although statistically there were no differences. We concluded that the modified air-dry method provided a larger source of mitotic nuclei for chromosome analyses of cloned bovine embryos.

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