Expression Of GDF9 Gene In The Reproductive Organ Of Female Sheep And Its Regulation Of Lambing Rate

Growth differentiation factor 9(GDF9)is one of the growth factors controlling follicular development and ovulation efficiency in animals.Therefore,GDF9 gene is considered as one of the most important genes controlling the reproductive rate of animals.Raising lambing rate has been the goal for sheep industry since most sheep breeds are uniparousfowl estrus seasonally.In this paper,small-tailed Han sheep of 2 months(before sexual maturity),6 months(sexual maturity)and 12 months(after sexual maturation)were chosen for study,and thetranscriptional and protein expression of ovary,fallopian tubes and endometrium were tested to describe its regulatory role in the development of female genitalia.After that,PCR-SSCP was used to detect the SNPs of GDF9 gene in broiler Merino sheep with double-lamb and single-lamb,thus to verify the relationship between the gene and the little size of sheep and possibility of GDF9 gene for candidate.The results are as follows:1.Immunohistochemistry was used to study the protein localization of GDF9 protein in the ovaries,fallopian tubes,endometrium and other major reproductive organs of female Han sheep at different ages.The results showed that GDF9 protein can be expressed in the endometrium,fallopian tubes and ovary of 3 different age groups at 2 months,6 months and 12 months of age.2.RT-PCR was used to detect mRNA expression and difference of GDF9 gene in ovary,fallopian tube,endometrium and other reproductive organs in 3 different ages of female small-tailed Han sheep.The results showed that mRNA of GDF9 gene was expressed in all reproductive organs of3 age groups,and there were differences among organs and ages.(1)The mRNA expression of GDF9 gene was significantly different among the ovary,fallopian tube and endometrium of 2-month-old female small-tailed Han sheep(P<0.01);there was no significant difference in the mRNA expression of GDF9 gene between the endometrium and fallopian tube of 6-month-old female small-tailed Han sheep(P>0.05),and there was significant difference between the endometrium and the ovary of 6-month-old female small-tailed Han sheep(P<0.05).There was an extremely significant difference between the expression of fallopian tube and the ovary(P<0.01);there was no significant difference in the mRNA expression of GDF9 gene between the fallopian tube and uterus,the uterus and the ovary of the 12-month-old Han sheep(P>0.05),but there was a significant difference between the fallopian tubes and the ovary(P<0.01).(2)There was no significant difference in the mRNA expression of GDF9 gene in endometrium between 2 months and 6 months of age(P>0.05);However,there was a significant difference between 2 months and 12 months,6 months and 12 months of age(P<0.05);there was no significant difference in the mRNA expression of GDF9 gene in the fallopian tube of the three ages(P>0.05);there was a significant difference in the mRNA expression ofGDF9 gene in the ovary between the 2 months and 6 months of age,2 months and 12 months of age(P<0.01),but there was no significant difference between 6 months and 12 months of age(P>0.05).3.PCR-SSCP method was used tostudy single nucleotide polymorphisms of the GDF9 gene in single-lamb,double-lambQianhua mutton merino sheep and polytocous female small-tailed Han sheep.The results showed that two genotypes such as AA and AB were detected in all three groups.The allele frequencies of A were 0.7,0.9,and 0.925.The allele frequencies of B were 0.3,0.1,and 0.075.Sequencing results showed that a single base mutation of G?A occurred at the 260 bp of exon 1 of type AB.Based on the differences in genotypic distribution,the mutation rate of single-lamb female Qianhua mutton merino sheep was significantly higher than that of double-lamb(P<0.05);The mutation rate of the single-lamb femaleQianhua mutton merino sheep was extremely significantly higher than that of polytocous female small-tailed Han sheep(P<0.01);there was no significant difference in the mutation rate between double-lamb femaleQianhua mutton merino sheep and polytocous female small-tailed Han sheep(P>0.05).The GDF9 gene could be used as candidate gene of sheep.