Canine cloning have been succeeded for a decade. To obtain in vivo matured dog oocytes, Serum progesterone (P4) level were employed for ovulate determination. However, accuracy of P4 methods is not satisfied. The aim of this study was to compare both methods of serum estradiol (E2) and P4 on the accuracy of canine ovulation determination. Canine serum P4 and E2 concentration during both proestrus and estrus were detected. Correlation between accuracy of each method and environment temperature were analyzed. Following ovulation, oocytes were collected by surgery. As a result, higher percentage of mature oocytes was obtained when using E2 (56.43%) as compared to P4 (39.60%). Accuracy of P4 increased from spring (30.76%) to summer (47.92%) and decreased in autumn (37.50%) and winter (29.16%) gradually. Especially, E2 maintained about 50% to 65% whatever the season and temperature. Correlation analyze showed that dynamic of P4 accuracy highly correlated with environment temperate (Rp4=0.862) but E2 could not be affected by the temperature (RE2=0.199). To determine whether obtained oocytes by E2 method could be used for canine cloning, twenty canines were selected as oocyte donors, and two puppies were produced after somatic cell nuclear transfer(SCNT) and embryo transfer(ET) with the oocytes by E2 method. In conclusion, comparing to the P4 method, the E2 is an accuracy and reliable method for canine cloning.

To obtain in vivo matured oocytes for dog cloning, serum progesterone (P4) level were employed for ovulate determination. Radioactive immunoassay (RIA) is a traditional serum hormone assay method with highly radioactivity. The aim of this study was to evaluate the reliability of RIA and to compare its canine serum P4 concentration determination accuracy to that of the electric chemiluminescence immunoassay (ECLI). To obtain in vivo matured oocytes for canine somatic cell nuclear transfer, serum P4 levels were accurately measured with both methods of RIA and ECLI. Although both methods detected similar P4 level before ovulation, the mean P4 concentration using ECLI was significantly higher than that using RIA from 3days before ovulation. Following ovulation, oocytes were collected by surgery, and a lower percentage of mature oocytes were observed using ECLI (39%) as compared to RIA (67%) if 4-8ng/ml of P4 were criteria for determination of ovulation. On other hand, high percentage of mature oocytes was observed using ECLI when 6–15 ng/mL of progesterone was criteria for ovulation determination. To determine whether in vivo oocytes obtained by ECLI method could be used for canine cloning, six canines were selected as oocyte donors and two puppies were produced after SCNT and embryo transfer. In conclusion, compared to the traditional RIA method, the ECLI method is a safe and reliable method for canine cloning.