| The Little-tailed Han Sheep is a kind of famous sheep line with multiplets and non-season oestrum in the world. Announcing the regulatory mechanism of multiplets on the level of molecular biology is the base to find the molecular mechanism of high fertility, use it in the breeding, and select the precise location on marker assistant selection (MAS). BMPR-IB and BMP15 as candidate genes on the fecundity of Little Tailed Han Sheep were studied and analyzed for their mutations and genetic effects. The results showed that there was a same mutation in BMPR-IB gene(A746G)of Little-tailed Han Sheep as that of Booroola Merino. The BB mutation genotype was superior in prolific Little Tailed Han Sheep, and the ewes with genotype BB had 0.97 (P<0.05)and 1.5(P<0.01)lambs more than those with genotype ++ in the first parity and later parities, respectively. It could be inferred that BMPR-IB gene was related with the major gene that controls the high prolificacy of Little-tailed Han Sheep. However, there was not mutation of V31D or Q23Ter in BMP15 gene of Little Tailed Han Sheep; it showed that the fecundity mechanism of Little Tailed Han Sheep was different from that of Romney sheep. Then it was ruled out the possibility that the ovulation of Little-tailed Han Sheep was affected by the mutation of BMP15 gene.The transcriptional promoter control region of 5' terminal in another gene of FSHR was cloned and analyzed that 15 regulatory elements of transcript region shown no difference among Little-tailed Han sheep, Tan sheep and sheep from Austrian to imply that the mutation of transcript region had no effects on transcription of FSHR. In this research, the cDNA library of ovary from Little-tailed Han sheep in the estrus was firstly constructed by SMART (switching mechanism at 5' end of RNA transcript) approach in the world. The plaque titer of this library was 1(109pfu/ml, the recombinant percentage was 96%, and the fragment length of inserted average cDNA was 1.5kb. Choosing 380 samples randomly to sequence, we get 338 ESTs by ligation. These ESTs were classed into 191 contig, among which the number of more than 2kb size was 7 and the number between 1~2kb size was 69. By clustering these gene contig, low expression of one copy were 148 of 77.49%, moderate expression of 2 to 5 copies were 33 of 17.28% and high expression of more than 5 copies were 12 of 6.28%. In the comparison, we had found 89 new ESTs of 191 ESTs which had no homology in the GenBank and EMBL, 88 ESTs had the homologies in the species of Human, Mouse, Bovine and 9 ESTs in the sheep, and 14 ESTs had unknown function. A method of mRNA Differential display PCR (DDRT-PCR) is firstly used in this paper to identify the differentially expressed genes infecting ovulation rate between the little-tailed sheep and Tan sheep. To improve further the efficiency and reproducibility of this method, this report systematically examines six critical parameters of standard DDRT-PCR by positive-cross test. The experimental findings delineated the best possible DDRT-PCR conditions to get 24 special bands by 3 anchor primers and 8 random primers to consist of 24 pair primers and reverse Northern hybridization to decrease the false positive bands. By comparison on BLAST website, 5 ESTs had one copy and 13 ESTs had several copy. Among these 18 ESTs, 6 ESTs cound find the homology gene with CDS and sure function, 3 ESTs can find homology gene but no CDS, and 9 ESTs had not homology gene. To further analysis the ESTs, we could find homology gene of NOS2,tensin,TCRA ,CDKN1A ,ESR1,ACTB which expression specially in the Little-tailed Sheep, but the relation of these genes with ovulation rate remained further study.As mentioned above, DD-PCR was also performed for mRNA expression of differential gene in the different development phase of follicles. The size larger than 5mm diameter choose from the delaminated antral follicles were contributed into the large group, while the size smaller than 3 mm diameter were contributed into the small group. By differential expressi...
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