【作者】 陶勇；

【导师】 王金玉；

【作者基本信息】 扬州大学 ， 动物遗传育种与繁殖， 2008， 博士

【摘要】 促黑激素皮质受体4(Melanocortin-4 receptor,MC4R)是动物下丘脑腹内侧核分泌的一类肽类物质,参与抑制体重的增加。垂体特异性转录因子(pituitary specifictranscription factor 1,POU1F1)参与调节机体的细胞生长与发育,促使生长激素(GH)、催乳素(PRL)和促甲状腺释放激素Beta因子(TSH-β)等基因的表达。鉴于MC4R基因和POU1F1基因对动物生长发育过程中的重要作用,本研究以优质肉鸡品种京海黄鸡为试验材料,采用PCR-SSCP研究了MC4R和POU1F1基因的单核苷酸多态性,并与该品种的生长及屠宰性状进行联合分析,为京海黄鸡分子标记辅助选择提供有效的遗传标记。同时,采用半定量RT-PCR技术分析比较了MC4R和POU1F1及相关基因在京海黄鸡和尤溪麻鸡中表达水平及相关关系,并与屠宰性状进行了相关分析。研究结果如下:(1)MC4R基因引物P1在京海黄鸡中检测到AA、AB、BB三种基因型,A、B等位基因频率分别为0.929和0.071;引物P2检测到CC、CD、DD三种基因型,C、D等位基因频率分别为0.72和0.28。卡方检验表明在这两个基因位点上,基因频率和基因型频率都未达到Hardy-Weinberg平衡状态。测序结果表明,引物P1在第662bp处发生了G→C的突变,引物P2多态位点是由于DNA的733-734bp之间插入一个C碱基引起的。这两个多态位点均为在鸡MC4R基因研究中首次被检测到。(2)POU1F1基因引物P1在京海黄鸡中检测到AA、AB、BB三种基因型,A、B等位基因频率分别为0.925和0.075;引物P6检测到AA、AB、BB三种基因型,A、B等位基因频率分别为0.5和0.5;引物P11检测到AA、AB、BB三种基因型,A、B等位基因频率分别为0.396和0.604。卡方检验表明,引物P1检测到的基因位点上,等位基因频率和基因型频率未达到Hardy-Weinberg平衡状态,而引物P6、P11的基因位点均达到Hardy-Weinberg平衡状态。测序结果发现,引物P1在第1876bp处发生了G→A突变,1882bp处发生C→T突变;引物P6在3684bp处发生了A→T的突变;引物P11在4573bp处发生了T→C的突变。(3)MC4R基因P1引物AB基因型个体、P2引物DD基因型个体的体重、体尺及屠宰性能均略高于其他基因型,组合基因型AA/DD的体重、体尺要好于其他组合。POU1F1基因P1引物AA基因型个体、P6引物AA型个体、P11引物AB型个体的的体重、体尺及屠宰性能均略高于其他基因型,P1、P6组合AA/AB,P1、P11组合AA/AB,P、P11组合AB/AB效应最佳。因此,MC4R及POU1F1基因的SNPs多态位点对京海黄鸡的生长及屠宰性状的影响均比较重要,可以尝试将其作为相关性状的分子标记用于鸡的分子标记辅助选择。(4)不同基因型内个体体尺性状、屠宰性能间相关关系分析表明,优势基因型个体的体斜长、胸骨长、胸宽、胫长、胴体重、半净膛重、全净膛重、胸肌重、腿肌重等指标间绝大部分存在着显著或极显著的相关关系,但与肌胃重之间一般没有显著的相关关系。(5)京海黄鸡POU1F1、GH、PRL基因的表达量显著地高于尤溪麻鸡,京海黄鸡公鸡MC4R基因的表达量显著高于尤溪麻鸡公鸡,母鸡及总体间差异不显著。京海黄鸡与尤溪麻鸡群体内,POU1F1与GH、PRL基因的表达水平间存在着极显著的正相关关系。(6)京海黄鸡群体内,MC4R基因表达量与肝重极显著相关;尤溪麻鸡群体内,MC4R基因表达量与肝重、心重极显著相关。更多还原

【Abstract】 Melanocortin-4 receptor(MC4R) is a kind of peptide secreted by animal ventral hypothalamus nuclear,involved in inhibiting the increase in body weight.Pituitary specific transcription factor 1(POU1F1) regulated the growth and development of body cell,fasted the gene expression such as GH,PRL,TSH-β.In order to give the effective molecular markers for the marker-assisted selection(MAS) on early production performance of chickens,SNPs polymorphism of MC4R and POU1F1 gene were conducted on Jinghai Yellow chicken breed by PCR-SSCP and DNA sequencing method. The relationship between SNPs polymorphism and growth performance,carcass characteristics had also been analyzed by GLM model.The differential expression level were detected about POU1F1 and relative gene in pituitary,and MC4R gene in adrenal gland,and compared between Jinghai Yellow chicken and Youxima chicken.The results were as follows:(1)Three genotypes of AA,AB,AB with primer P1 of MC4R gene were detected,the allele frequency of A,B were 0.929 and 0.071 respectively.With primer P2,results showed three genotypes of CC,CD,DD,and the allele frequency of C and D allele was 0.72 and 0.28 respectively.The gene frequencies and genotype frequencies both reached the Hardy-Weinberg equilibrium at those two loci indicated by the x~2 test.Through sequencing alignment,results showed that there was a G→C on the site of 662bp with primer P1 of MC4R gene,and a insertion of C base between the site of 733 and 734bp. These two polymorphism sites were both detected first in MC4R gene of chicken.(2)Three genotypes of AA,AB,AB with primer P1of POU1F1 gene were detected, the allele frequency of A,B were 0.925 and 0.075 respectively.With primer P6,showed three genotypes of AA,AB,AB,and the allele frequency of A and B allele was 0.5 and 0.5 respectively.With primer P11,showed three genotypes of AA,AB,AB,and the allele frequency of A and B allele was 0.396 and 0.604 respectively.The gene frequencies and genotype frequencies both reached the Hardy-Weinberg equilibrium at P6,P11 loci indicated by the x~2 test,but P1 loci not reached.Through sequencing alignment,results showed that there was a G→C on the site of 1876 bp,and a C→T on the site of 1882 bp with primer P1 of POU1F1 gene,and a A→T on the site of 3684 bp with primer P6,a T→C on the site of 4573 bp with primer P11.(3)The body weight,body measure and carcass trait of AB genotype individual of P1 primer in MC4R gene were better than other genotype,DD genotype individual was the same.The body weight and measure of the combined genotype AA/DD individual were better than other combination.That was the same such as AA genotype of P1,AA genotype of P6,AB genotype of P11 in POU1F1 gene.So,Those SNPs of MC4R and POU1F1 gene had important effect on the growth performance and carcass characteristics in Jinghai Yellow chicken breed,and could be used to MAS as molecular markers of correlation characteristics.(4)The correlation between body measure trait and carcass were analyzed.The results showed that there was a significant or very significant correlation between the traits of advantage genotype.But there was not significant correlation between the muscle stomach and other traits.(5)The expression level of POU1F1、GH、PRL gene in pituitary of Jinghai Yellow chicken had significantly higher than it in Youxima Chicken.The expression level of MC4R gene in adrenal gland of Jinghai Yellow male chicken had significantly higher than it in Youxima male Chicken,but it was not significant both in female and total.Both in Jinghai Yellow chicken and Youxima chicken,the expression level had a positive correlation between POU1F1 gene and GH,PRL gene.(6)In Jinghai Yellow chicken,the expression level of MC4R gene had a greatly significant correlation with liver weight;but in Youxima chicken,the expression level of MC4R gene had a greatly significant correlation with liver weight and heart weight.更多还原