Partial Sequence Study On The Follicle-stimulating Hormone And Luteinizing Hormone Beta Subunit Gene Of Goat

It is the man's hopeful object to reduce production cost and improve economic benefit by increasing the birth rate of pig, bovine, sheep and goat. Therefore, study on major gene influencing litter size at genetic structure and gene expression has been paid a great attention to in the world wide range. FSH and LH are critical hormones to follicular growth, development, dominance, maturation and ovulation. They also influence the growth of the reproductive system and the quality of sperm. An inserted fragment of FSHp was considered as the QTL of litter size in pig, but little things about goat FSHβ and LHβ gene was known except for FSHβ mRNA. In this study, Comparing the FSHβ and LHβ gene of sheep, bovine and pig in GenBank, the primers were designed, amplified and the sequenced the FSHp intronl, intron2, ex-3 and LHβ gene of 21 goat breeds (including 15 native breeds, 2 developed breeds and 4 introduced breeds) at first time, Studied on their genetic structure, base mutations within and among breeds and species, and phylogeny analysis.The complete length of goat FSHβ intronl was 641bp, the content of A, C, G and T was 35.9%, 13.9%, 17.3% and 32.9%, respectively. AT base (68.8%) was significantly higher than the C+G (31.2%) .There were only two mutations in this region, one was 71 loci (A-G), the other was 289(A-G).So the goat group had only two different haplotypes, Angora goat(An), Liaoning Cashmere goat(L), Jianyang native goat(N), Lezhi black goat (Lz) and Nubian goat(Nb)shared a haplotype, and others had the other one.In the 1560bp sequence of FSHp intron2, the content of A, T, C, G was 35.7%, 32.8%, 16.8%, and 14.8%, respectively. There were 14 bases deletion and 27 mutation loci, the number of Singleton variable sites (two variants) was 7, and Parsimony informative sites (two variants) 20, only one was three variants (890 loci A-G-T).The transitionsal pairs(Si) and transversional pairs(Sv) were equal. A-G (8 times & 13 total) was the main style of transition, and the T-G (6 times & 13 total) and A-T (4 & 13 total) were the main of transversion. Many breeds had its unique haplotypes, such as Tibet goat (Zh), LiaoNing Cashmere goat(L), and Boer goat(Bo), et al. the goat breeds shared many haplotypes except for JianYang Daer goat (Jy) and Sannen dairy goat (Sa).The partial sequence of FSHp exon3 was studied, the 222bp of 574bp at the beginning was coding, the other was non-coding. No mutation exited in 3' flanking region within or between breeds, while there were 6 mutations in the coding region, 5 of them leaded to amino acid change, the details were below: a59g/Y76C, al42g/K104E, g89t/R86L, tl65c/R115Q, and al87c /P119T, R86L was not only in the region of FSHP subunit binding to a, but FSH binding to its receptor. At the 3rd site of codon, GC content amounted to 68.7%, and the codon with G or C at 3rd site was more useful.The total sequencing length of goat LHP was 929bp, including 5'UTR(136bp, Partial), exonl(26bp), intronl(297bp), exon2(168bp), intron2 (235bp), and partial sequence of exon3(67bp).The content of each base was 17.6%(T), 28.1%(C), 17.1%(A) and 37.2%(A), respectively. LHp was rich GC region, because LHP showed high level of G+C (65.3%) content, especially the third site of codon, amount to 89%. There were 9 polymorphism sites in goat group, two singleton variable sites (two variants), seven Parsimony informative sites (two variants), one site had three variants. Three mutation loci in the 5'UTR, 3 loci in the intronl, 2 loci in exon2, and 1 loci in exon3, they did not change the amino acid. The coding sequence presented73 amino acids, and the synonymous codon usage bias was so significant.There were 14 promoter elements or regulating elements existed by scanning the 5'flanking region of LHP gene, three Sp-1 elements, two Egr-1 and GSE(SF-l), respectively, the TATA box, GC box, AP-1 element, Runx factors, CAC box, LYF-1 and Ptx-1 was only one each. The NF-Y, one of the house-keeping gene basic elements, was not found, maybe the 5'UTR sequence was too short in this study. There were two mutations in the 5'flanking region exiting in the 5'Egr-l element among goat breeds.The polymorphism of Nanjiang-Huang-Goat LH(3 region studied using restriction endonuclease Sph I by PCR-RFLP. 2 alleles, A and B, and 3 genotypes, AA(4.065%), AB(44.309%), BB(52.606%) were identified. The AA was caused by two mutations of loci 401, so it had very low frequency (4.065%). But the litter size and birth weight of litter had no significant difference between different genotypes, so the mutation of 401 loci had not change the two properties directly.Though the FSHP and LHP gene was very stable, the phylogenetic trees showed that all haplotypes were divided into multiple obvious clusters. Some goat breeds had special base mutation, so it was important to protect the diversity of different breeds.Regardless of the intron or exon, and the sequence length more than 700bp or not, they could be used to construct phylogenetic trees among species. While analyzing the evolution between species, be careful about the GC difference between the close relative species especially, or you would not get correct results.