【作者】 侯启瑞；

【导师】 王金玉；

【作者基本信息】 扬州大学 ， 动物遗传育种与繁殖， 2010， 博士

【摘要】 以优质肉鸡新品种京海黄鸡为试验材料,利用3个生长曲线模型和4个产蛋曲线模型拟合京海黄鸡生长曲线和产蛋率曲线,分析该品种的生长和产蛋规律;利用PCR-SSCP技术检测IGF1和IGF2基因不同区段的SNPs,首次检测LYZ基因外显子和EAV-HP DNA序列的SNPs,并分析所得SNPs与京海黄鸡部分经济性状之间的关系,为京海黄鸡分子标记辅助选择提供有效的遗传标记;在对单基因标记研究的基础上,对IGF1&IGF2基因和LYZ&gag-env基因进行互作效应分析,合并基因型,对京海黄鸡重要经济性状进行多基因分子标记联合分析,以期为该品种筛选出更准确的遗传标记。主要研究结果如下:⑴京海黄鸡累积生长曲线呈S型,Logistic、Bertalanfy和Gompertz三种模型均能很好的拟合京海黄鸡生长曲线,其中Gompertz模型拟合的效果最好;杨宁模型和分室模型均能很好的拟合京海黄鸡产蛋率曲线,其中杨宁模型拟合度较高,比较适合拟合京海黄鸡的产蛋率曲线。⑵鸡IGF1基因外显子1、2和4未检测到突变,外显子3和5’非编码区存在突变位点,且外显子3属于沉默突变,该突变没有引起氨基酸序列的改变;IGF1基因基因型分布处于Hardy-Weinberg平衡状态,说明京海黄鸡群体遗传性比较稳定,该品种仍具有较大的选育潜力。⑶IGF1基因对京海黄鸡生长性状有重要影响,可以作为遗传标记对鸡的相关性状进行分子标记辅助选择;IGF1基因可能与繁殖数量性状位点有一定的距离,因而与繁殖性状之间的关系可能随着种群、品系和家系的不同而不同。⑷鸡IGF2基因外显子1、2和3均存在突变位点,IGF2基因外显子1和2碱基突变改变了氨基酸序列,外显子3突变属于沉默突变;IGF2基因2对引物基因型分布处于Hardy-Weinberg平衡状态,1对引物基因型分布极不平衡,说明该群体在对某些性状的选育过程中影响到了该基因的随机遗传。⑸IGF2基因对鸡生长性状的影响在不同品种中不同生长发育阶段表现不同。初步推断IGF2基因可能不是鸡繁殖性状的主效基因,但可能与繁殖性状数量位点有一定的连锁关系,在鸡的性成熟、产蛋量和蛋重方面有影响。⑹首次对鸡LYZ基因外显子区域SNPs进行扫描,发现外显子1和2存在同义突变,外显子3和4未检测到突变位点;LYZ基因外显子1基因型分布从不平衡到平衡可能与京海黄鸡的人工选择有关,这也正说明该基因与京海黄鸡重要经济性状有很大关联,外显子2检测到5种基因型,揭示出该群体的遗传多样性十分丰富。⑺鸡LYZ基因与京海黄鸡蛋清中溶菌酶的含量和活力没有显著相关,表示该基因对溶菌酶在蛋清中的表达没有显著影响;通过对京海黄鸡J+和J- 2个品系的研究,首次发现该基因与部分经济性状显著相关,初步推断LYZ基因可能是一个调控鸡生长发育的主基因或与主基因紧密连锁。⑻建立了测定蛋清中溶菌酶含量和活力的标准方法,为家禽蛋清中溶菌酶的测定提供了一套操作性强、易于掌握、结果可靠稳定的途径。⑼首次对鸡EAV-HP DNA序列部分区域进行SNPs扫描,共发现7个多态位点,其中2个位点为沉默突变,5个位点突变造成了氨基酸序列的改变,改变可能会使蛋白结构和生物学功能产生差异,直接或间接影响动物机体的生长发育;京海黄鸡在品系繁育过程中经过了多重人工选择,该基因所处的位置在品系选育目标对应的数量遗传区域内或邻近区域,对两个不同品系所施加的人工选择压无意间改变了基因的随机遗传规律,使基因型在群体中的分布极不平衡。⑽通过基因型&单倍型与京海黄鸡两个品系生长性状的相关性分析,初步认为EAV-HP DNA序列中的gag-env融合基因与鸡生长性状显著相关,且对京海黄鸡两个品系的作用方向不同。⑾京海黄鸡IGF1基因和IGF2基因对0周龄体重、12周龄体重和开产日龄均有互作效应,不同基因型组合间差异显著;LYZ基因和gag-env基因对京海黄鸡8、12和16周龄体重存在互作效应,不同基因型组合间差异显著。⑿单基因基因型分析结果与两基因基因型组合分析结果不完全一样,利用影响经济性状的两个基因的SNPs,甚至更多基因的SNPs进行分子标记辅助育种,比单个基因进行选择的风险小。在京海黄鸡生产上,选留I3* IG1 AA/CC型个体可提高出生重,选留I6*IG1 CC/BB型个体可提高12周龄体重,剔除I6*IG1 DD/BB型个体可减小开产日龄,选留L1*EA6 GA/AC和GA/CC个体可提高12和16周龄体重,选留L2*EA3 TN/AC和L2*EA6 CC/AA、CC/BB、CC/CC、CT/AC个体可提高8周龄体重。更多还原

【Abstract】 Use of three growth curve models and four egg production curve modles analysed Jinghai yellow chicken growth and laying traits laws. In the first part of the present study, we chose IGF1, IGF2, LYZ genes and EAV-HP DNA sequence as candidate genes for economic traits in Jinghai yellow chicken. To explore the relationship between these genes with economic traits in chicken, single strand conformation (SSCP) analysis and sequencing reactions were performed to screen single nucleotide polymorphisms (SNPs) in Jinghai yellow chicken. In the second part, we analyzed the effects of combined genotypes of different genes involved in IGF1*IGF2 and LYZ*gag-env interaction modes. The main results were as follows:⑴The cululative weight curve of Jinghai yellow chicken was like an“S”line. Logistic, Bertalanfy and Gompertz models well fitted the growth curve of Jinghai yellow chicken, and the Gompertz model fitted best. Yang Ning and McMillan models fitted well in Jinghai yellow chicken egg production curve, and Yang Ning model better.⑵No mutations were detected in exon 1, 2 and 4 of Chicken IGF1gene. Mutations were found in exon 3 and 5’non-coding regions. The mutation in exon 3 was silent mutation, didn’t cause changes in amino aci sequence. Genotype ditributions of IGF1 gene were in Hardy-Weinberg equilibrium, indicating the genetic nature of Jinghai yellow chicken population was relatively stable. The species still had great patential for breeding.⑶The chicken IGF1 gene had a major impact on growth traits of Jinghai yellow chicken, which could be used as a genetic marker in molecular marker-assisted selection on ralated traits. IGF1 gene may be linked with reproductive QTL to a certain distance, thus the relationship with reproductive traits may change with different species, strains and families.⑷Mutations were detected in exon 1, 2 and 3 of chicken IGF2 gene. The mutations of exon 1 and 2 changed amino acid seqence, excluding exon 3 silent. Distribution of genotypes produced by two pairs of primers was in Hardy-Weinberg equilibrium, while by one pair of primers was extremely uneven. This indicated that the seletion process of certain traits affected random genetic of the gene in Jinghai yellow chiken.⑸The effect of IGF2 gene on growth traits differ at different growth stages in different strains of chicken. To be concluded that chicken IGF2 gene might not be the major gene controlling reproductive traits, but might be in a certain linkage with reproductive traits. The chicken IGF2 gene effected sexual maturity, egg producton and egg weight in Jinghai yellow chicken.⑹By SNPs scanning in exon region of chicken LYZ gene, synonymous mutations were detected in exon 1 and 2, excluding in exon 3 and 4. The distribution of genotypes in exon 1 changed from imbalance to balance might be related to artificial selection, which indicated that the chicken LYZ gene had great relevance to important economic traits. Five genotypes were found in exon 2, revealing the genetic diversity was very rich.⑺The chicken LYZ gene did not significantly associated with lysozyme content and enzyme activity in egg white, indicating that the gene had no significant effect on lysozyme expression in egg. The gene was found in significantly relationship with some economic traits of chicken for the first time by study on Jinghai yellow chicken J+ and J- lines. We initially infered that the LYZ gene might be a major gene or closely linked with major gene regulating chicken’growth and development.⑻To provide a feasible, easy, reliable and stable way, a standard method for determining poultry egg white lysozyme content and enzyme activity was established.⑼By scanning for the first time in part of the chicken EAV-HP DNA sequence, SNPs were found in more than seven polymorphic sites, of which two sites as silent mutations, five mutations caused changes in amino acid sequence. Amino acid changes might make protein be different in structure and biological function, and directly of indirectly affected the growth of animal body. The J+ and J- lines of Jinghai yellow chicken were bred by multiple artificial selection process. The location of the gene might be in the target QTL genetic region or adjacent area. Artificial selection pressure imposed on the two different strains inadvertently changed the random inheritance of the gene, so that the distribution of genotypes in the population was extremly uneven.⑽By correlation analysis of genotype&haplotype and growth traits of two strains of Jinghai yellow chicken, We concluded that the gag-env fusion gene of EAV-HP significantly related with chicken growth traits, and the direction of effects differred in J+ and J- strains.⑾Chicken IGF1 and IGF2 genes had interaction on 0-week, 12-week BW and age of first egg in Jinghai yellow chicken. The economic traits of chicken with different genotype combinations were significantly different. Chicken LYZ and gag-env genes had interaction on 8-week, 12-week and 16-week BW in Jinghai yellow chicken.⑿The analysis of single-gene genotype combiantion and two genes with growth traits were not exactly the same. Using SNPs of two genes or more genes for marker-assisted breeding had less risk than using SNPs of a single gene. On Jinghai yellow chicken breeding, selecting I3*IG1 AA/CC individuals could improve birth weight, choosing I6*IG1 CC/BB chicken could increase 12-week BW, excluding I6*IG1 DD/BB individuals could lead to early age of fist egg, saving L1*EA6 GA/AC and GA/CC chicken could increas 12-week and 16-week BW, reserving L2*EA3 TN/AC and L2*EA6 CC/AA, CC/BB, CC/CC, CT/AC individuals could increase 8-week BW.更多还原