Analysis of specific molecular markers
Analysis of specific molecular markers and important genes in PGC from White Leghorn chicken
Su Jiun Deng and Jen-Wen Shiau
The purpose of this study is to investigate the developmental and pluripotentcy related genes expression profile of PGC obtained from day 5.5 and day 8.5 chick embryos after incubation. PGC was obtained by micromanipulation and the total RNA was purified by commercial kit. The cDNA was synthesized using RT-PCR with total RNA as a template. The genes expression was detected by real-time PCR using various kinds of primers and the results showed that all the genes detected were expression both in day 5.5 and day 8.5 chick embryos except the Blimpl. Some genes that selected from the chicken germ cell database were also showed expression both in day 5.5 and day 8.5 chick embryos. Cvh is an important gene and preferentially expressed in whole period of germ cell differentiation. We constructured the siRNA of Cvh and transfect to PGCs. After 24 hours of transfection, the expression of Cvh in male PGCs was down-regulated, but not in female's PGCs. Then we compared male and female specific gene expression after siRNA treatment. Pouv up-regulated in both sex, indicated the PGCs prefer to stay in PGCs but germ cells. Sox2, Sox9 also up-regulated lightly in male PGCs. In female PGCs, LhX8 was down-regulated by siRNA, which is the important factor about maintain the oocyte and differentiated on early oogenesis. Our findings show that Cvh is a critical factor for maintenance and differentiation of the oocyte during early oogenesis, and it acts in part by down-regulating the Lhx8 pathway.