科研之友
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摘要
The objective of this study was to identify the sex of in vitro produced buffalo embryos using an efficient polymerase chain reaction (PCR) assay. Buffalo oocytes collected from slaughtered animals were in vitro matured, fertilized and cultured for development until morula and blastocyst stage. The embryos were washed twice in phosphate buffered saline (PBS) and once in 1X PCR buffer then each embryo was placed separately in 20 mu l 1X PCR buffer and stored at -20 degrees C for DNA extraction and PCR assay. A pair of bovine satellite primers common to both male and female and a pair of male-specific primers (BRY.1) which targeted male-specific sequence in the buffalo DNA were used in PCR assay. The assay was carried out on whole blood samples, collected from adult 10 male and 10 female total 20 buffaloes and 50 morula, 50 blastocyst in vitro fertilized buffalo embryos. When DNA samples from blood were amplified, the sex was determined by PCR always corresponded to the anatomical sex. The results are females representing only the 216 bp while males representing both 300 and 216 bp. The percentage of sex determination is 20 out of 20 (100%) accurate for blood samples. The morula and blastocyst stage sex determination of 44 out of 50 (88%) and 46 out of 50 (92%) respectively, were successfully determination using the multiplex PCR assay. In conclusion, this multiplex PCR assay could be used as a reliable and efficient tool for sex determination of in vitro fertilized buffalo embryos satisfactory.
- 出版日期2013
