Poymorphism Analysis Of Goat TRα And CRABPⅡ Gene In Three Goat Breeds And Its Expression In Cashmere Goat Fetus Skin

Using Inner Mongolia Cashmere goat as experimental materials, we cloned and sequenced TRa and CRABPⅡ genes. Real-time RT-PCR method was used to quantify the mRNA expression of TRa and CRABPⅡ genes in skin from Inner Mongolia Cashmere goat embryos at different development stages. In addition, PCR-RFLP method was used to detect the polymorphisms of TRa and CRABPⅡ genes in Inner Mongolia Cashmere goat, Nanjiang yellow goat and Tibet goat. The main results are reported as follows.(1) The CDS of goat TRa and CRABPⅡ were1233bp and417bp, which encoded410and138amino acid residues respectively. Comparison of CDS sequences of goat TRa with those of sheep, cow, pig, huamn and mouse, the similarity were more than95%, but the highter similarity(99%) presented between sheep and cow. It also showed that the similarity of amino acid sequences between the same species were more than99%. Comparison of CDS sequences of goat CRABPⅡ with those of cow, pig, huamn and mouse, the similarity were more than88%, but the highter similarity(98%) presented in cow.And it also showed that the similarity of amino acid sequences between the same species were more than93%.(2) The bioinformatics analysis of amino acids indicated that the goat TRa contains19phosphorylation sites, but not transmembrane domain, signal peptide and hydrophobic region. Its secondary structure includes49.27%of alpha helix,43.90%of random coil and6.83%of extended strand. The goat CRABP Ⅱ contains one conserver domain-lipocalin, two hydrophobic region,3phosphorylation sites. Its secondary structure includes41.30%of extended strand,33.33%of random coil and25.36%of alpha helix.(3) The expression level of TRa and CRABPⅡ gene in skin of Inner Mongolia Cashmere goat fetus had been detected at different days. The data showed that TRa gene was highest expressed at70d, reached the bottom at90d, and the difference was highly significant (P<0.01). CRABPⅡ gene was highest expressed at75d, reached the bottom at70d, and the difference was significant (P<0.05).(4) There was one mutation site found in intron1(A447G) of CRABPⅡ gene, and seven mutation sites found in intron6(C230T, G245A, C408T and A480G) and intron7 (A155G, G173A and G197C). Two genotypes (CC and CT) were obtained by analyzing C230T locus by technique of RFLP. Polymorphic analysis showed that this mutation was minuent polymorphic loci in the herd of Nanjiang yellow goat, Inner Mongolia Cashmere goat and Tibet goat, and it was in Hardy-Weinberg equilibrium in the three herds. G173A locus had three genotypes (GG、GA and AA) by technique of RFLP, but AA genotype was only existed in Tibet goat. And its analysises of polymorphic and Hardy-Weinberg equilibrium were as the same as the results of C230T locus.(5) The association analysis between each genotype of the restriction sites of two mutation sites of TRa gene and growth trait of Nanjiang yellow goat showed that there was no significant difference among the growth traits of different genotype of C230T locus and G173A locue, except that chest circumference of2-month goat in C230T locus was significant (P<0.05). The four combinations of genotype of C230T locus and G173A locus, which were CCGG (Ⅰ type), CCGA (Ⅱ type), CTGG (Ⅲ type), CTGA (Ⅳ type). The association analysis showed that except that withers height of one-year goat and body length of adult ewes between Ⅰ type and Ⅳ type was significant (P<0.05), there was no significant difference among other growth traits of different genotype combination.