Polymorphism And Expression Of ESR And FSHR Gene And Its Relations With Litter Size In Alopex Lagopus

The litter size of arctic fox is one of the important economic characters, increasing litter size can reduce the breeding number of fox, increase the output of the skin, bring huge economic benefits to fully; At the same time, improve the litter size is also the main goal of the arctic fox breeding work. Arctic foxes breed character is very low heritability of quantitative traits, it is difficult to get large genetic progress in the short term by the conventional breeding methods, and marker-assisted selection (MAS) provides an effective way. This study serves FSHR gene and ESR gene as candidate genes, cloning FSHR gene and ESR gene by PCR, and sequence analysis and homology comparison; detecting FSHR gene and ESR gene polymorphism by PCR-RFLP and PCR-SSCP technology sequence analysis of polymorphic loci, determining the mutation type, and polymorphic loci associated with litter size traits of arctic fox analysis; researching expression of different FSHR and ESR genotype of FSHR and ESR gene in ovarian tissue by realtime PCR technology and its effects on litter size traits. The results are as follows:(1) ESR gene sequence is1824bp, contains the start codon ATG, open reading frame encoding607amino acids, the sequence of A, T, Q C ratio were25.23%,22.69%,25.92%,26.16%, G+C content (52.08%) higher than that of the A+T content (47.92%). DNA binding area, there are eight conservative cysteine residues (C), constitute two zinc (zinc-C4), D-box (EGCKA), P-box (PATNQ), there is a signal DRNRRKS (aa244-250).(2)Arctic foxes FSHR gene sequence is1947bp and contains a start codon ATG, encoding648amino acids, the sequence of A, T, G, C ratio were25.17%,26.55%,21.42%,26.86%, G+C content (48.28%) is lower than the A+T content (51.72%).(3) the arctic fox ESR gene sequence of amino acids with chickens, sheep, cows, mice, pigs, people of homology respectively is74%,77%,87%,88%,75%,91%, arctic fox on evolutionary relationships are the closest animal relatives to pigs, and farthest to chickens, amino acid composition analysis showed that the highest levels is serine,tryptophan is the lowest.(4) FSHR gene sequence of amino acids with chickens, sheep, cows, mice, pigs, people of homology respectively is59%,73%,83%,83%,82%,92%, arctic fox on evolutionary relationships are the closest animal relatives to pigs, and farthest to chickens, amino acid composition analysis showed that the highest levels is serine, the tyrosine is minimum.(5)Arctic fox ESR protein in56～60,88～88,89～121,154～154,172～205,215～220,215～220,263～263,266～337,353～359,353～337,394～394,420～503,518～526,518～503amino acids exist15significant hydrophobic area and there is no cross membrane structure, amino acid sequence from196to196are ZnF_C4model sequence structure domain, the amino acid sequence from360to530is HOLI structure domain.(6)FSHR protein in5～16,25～33,288～296,304-309,332～332,338～349,357～366,357 ～349,402～402,417～456,485～485,553～572,587～600,587～600amino acids exist14significantly hydrophobic regions, amino acid sequence from367to389,402to424,402to466,490to512,490to554,575to597,575to629exist seven transmembrane structure.(7) the FSHR gene5’end of the wing of the arctic fox found no polymorphism; found polymorphism in the arctic fox FSHR gene exon10, detected three genotypes AA, AB, and BB, AA genotype is advantage in the arctic fox group, the frequency of A allele is0.82, the frequency of B is0.18; TNB and NBA of primiparous arctic foxes with BB genotype is1.08and1.58respectively than AA genotype (P<0.05), TNB and NBA of parous arctic foxes with BB genotype is1.72and1.03respectively than AA genotype (P<0.05).There is Câ†'T mutation at120bp in FSHR gene exon10.(8) ESR gene exon1of Arctic foxes exist polymorphism, A alleles is advantages, A allele frequency is0.68, the B allele frequency is0.32; TNB and NBA of primiparous arctic foxes with BB genotype is2.61and1.25respectively than AA genotype (P<0.05), TNB and NBA of parous arctic foxes with BB genotype is2.08and1.29respectively than AA genotype (P<0.05). There is Gâ†'C mutation at539bp in ESR gene exon1.(9)Combined genotype AA, AB, and BB show gradually increasing trend, primiparous TNB and NBA of genotype AAAA are7.42and6.22respectively, genetic type ABAB are7.46and7.07respectively, genotype BBBB are8.92and8.56respectively, parous TNB and NBA are also in the same rule.(10) FSHR mRNA level of BB genotype of the right ovary was significantly higher than that of AA genotype in the oestrum (P<0.01), BB genotype has significant difference with AA genotype in the left ovary (P<0.05)(11)ESR mRNA level of both ovary among the defferent genotypes have no significant difference (P>0.05).(12) FSHR mRNA of arctic foxes espresses during the different stage of estrous cycle in ovarian, the expression quantity of FSHR mRNA is on the rise from metaestrus to estrus, the highest expression quantity in the estrus,the lowest expression quantity in metaestrus.(13) FSHR positive cells distribute in the different stages of follicle of arctic foxes during the estrus cycle, FSHR positive cells mainly exist in the follicular cell and granulosa cell and theca cells and primary oocyte. From metaestrus to estrus, FSHR positive cells number is on the rise, the highest number in estrus, the lowest number in metaestrus. Number of positive cells increase with follicular development and increasing of follicular volume.