吐鲁番斗鸡攻击行为的表型与基因型分析

发布时间：2018-08-06 09:10

【摘要】：目的:通过“吐鲁番斗鸡攻击行为的表型与基因型分析”探讨吐鲁番斗鸡攻击行为表型和基因型的遗传特点,寻找可作为吐鲁番斗鸡攻击行为的分子标记,把分子标记辅助选择应用于吐鲁番斗鸡攻击行为的选育,为吐鲁番斗鸡攻击行为的分子育种奠定基础。方法:选择攻击性强的吐鲁番斗鸡与攻击性弱的新罗曼蛋鸡杂交产生F1代,采用荧光定量PCR技术确定SLC6A4基因与NR2E1基因在攻击性强的吐鲁番斗公鸡、攻击性弱的新罗曼蛋公鸡和F1代的大脑从出壳到性成熟不同时期阶段的表达变化;通过F1代鸡全同胞交配产生F2代,采集亲本、F1代和F2代三代四种类型鸡的血液样本,通过PCR-SSCP方法检测与攻击行为相关的SLC6A4基因与NR2E1基因全部外显子区和部分内含子区的SNP多态性,然后采用“镜像自斗法”和“配对打斗法”测定亲本、F1代的攻击行为表型,然后通过攻击行为表型与攻击行为相关的基因型之间进行方差分析验证SLC6A4基因和NR2E1基因与吐鲁番斗鸡攻击行为的关联性。结果:(1)通过对SLC6A4基因表达量在7个时期的分析发现其趋势成波浪形,0月龄、3月龄、6月龄都有较高的表达;在三种鸡群中不同时间段的表达量中发现SLC6A4的表达量与攻击行为呈正相关。NR2E1基因在7个时期的表达量呈逐渐升高趋势,且表达量均在6月龄达到最高;三种鸡群中不同时间段的表达量中发现NR2E1的表达量与攻击行为呈负相关。表明SLC6A4基因、NR2E1基因与吐鲁番斗鸡的攻击行为具有一定的相关,为后续的SLC6A4基因和NR2E1基因多态性分析奠定了理论基础,旨在获得SLC6A4基因和NR2E1基因的分子标记,为吐鲁番斗鸡的分子标记辅助选择提供参考依据。(2)在SLC6A4基因的外显子多态性检测中发现第一外显子有两个碱基突变位点(T6213768G、C6213750G)引起氨基酸的改变,且第一外显子的SNP位点所造成基因型的改变在吐鲁番斗鸡和新罗曼蛋鸡中有显著不同的优势等位基因和优势基因型。因此,进一步证实SLC6A4基因第一外显子的多态性可能与攻击行为有关,而在NR2E1基因的外显子多态性检测中未发现与攻击行为相关的基因多态性。(3)镜像打斗试验中,吐鲁番斗公鸡的直接攻击(Fighting)极显著高于杂交子一代公鸡(P0.01);而在非直接攻击(Non-fighting)上两者差异不显著。在配对打斗试验中,吐鲁番斗鸡的直接攻击(Fighting)显著高于杂交子一代(p0.05),然而二者之间在非直接攻击(Non-fighting)行为变量上的表现为差异不显著(P0.05)。(4)通过SLC6A4第一外显子的基因型与镜像和配对打斗测得表型的关联分析发现具有BB基因型的斗鸡与F1代的攻击性在镜像打斗试验中和配对打斗试验中均显著高于其它基因型的鸡,从而说明第一外显子的BB基因型与攻击行为有密切的相关性。结论:通过对攻击行为强的吐鲁番斗鸡、攻击行为弱的新罗曼蛋鸡及其杂交F1不同时期SLC6A4基因和NR2E1基因表达检测发现SLC6A4基因表达与攻击行为呈正相关,而NR2E1的基因表达则与攻击行为呈负相关;进而通过SLC6A4与NR2E1在吐鲁番斗鸡,新罗曼蛋鸡及F1、F2代的SNP多态性进行分析,在SLC6A4第一外显子内寻找到了与攻击行为相关的分子标记;通过吐鲁番斗鸡与F1代的表型检测发现两者之间在攻击性方面有显著的差异,进而通过攻击行为的表型与SLC6A4第一外显子的基因型关联分析发现BB基因型与攻击行为具有的相关性,可作为吐鲁番斗鸡分子辅助标记选择的参考基因型。
[Abstract]:Objective: To explore the genetic characteristics of the aggressive behavior phenotype and genotype of Turpan cockfighting chicken through the analysis of the phenotype and genotype of the aggressive behavior of Turpan chicken fighting, looking for the molecular markers that can be used as the attack behavior of the Turpan cockfighting, and applying the molecular marker assisted selection to the breeding of the attack behavior of the fighting chicken in Turpan, for the attack line of the fight chicken in Turpan. The basis for molecular breeding was established. Methods: the F1 generation was hybridized with aggressive Turpan fights and aggressive Xinluo man laying hens. Fluorescence quantitative PCR was used to determine the SLC6A4 and NR2E1 genes in aggressive Turpan cocks. The brains of the aggressive Xinluo man egg cock and the F1 generation were different from the shell to sexual maturity. The expression changes at the stage of the period; the blood samples of four types of chickens of the parents, the F1 generation and the three generation of the three generation of the F2 generation were collected through the whole sib mating of the chickens of the F2 generation, and the PCR-SSCP method was used to detect the SNP polymorphism of the SLC6A4 gene related to the attack behavior and all the exons and introns of the NR2E1 gene, and then the "mirror self fighting method" was used. "Paired fight method" measured the parents, the F1 generation of the aggressive behavior phenotype, and then analyzed the correlation between the SLC6A4 gene and the NR2E1 gene and the Turpan cockfight attack by the analysis of the variance between the aggressive behavior phenotype and the attack related genotypes. Results: (1) the expression of SLC6A4 gene expression was found in the 7 periods. The potential was wavy, 0 months, 3 month old, and 6 month old had higher expression. The expression of SLC6A4 was positively correlated with the attack behavior in the three chicken groups. The expression of.NR2E1 gene increased gradually in the 7 period, and the expression amount reached the highest in 6 month old. The expression of the three chicken groups at different time periods was the highest. It was found that the expression of NR2E1 was negatively correlated with the attack behavior, indicating that the SLC6A4 gene and the NR2E1 gene were related to the attack behavior of the Turpan cockfighting, which laid a theoretical basis for the subsequent SLC6A4 and NR2E1 gene polymorphism analysis, aiming at obtaining the molecular markers of the SLC6A4 gene and the NR2E1 base, as the molecular marker of the Turpan fighting chicken. 2. (2) in the exon polymorphism detection of the SLC6A4 gene, it was found that two base mutation sites (T6213768G, C6213750G) in exon of the first exon cause the change of amino acids, and the genotype changes caused by the SNP loci of the first exon have significant different advantages in the Turpan cockfighting and the Xinluo man laying hens. Therefore, it was further confirmed that the polymorphism of the first exon of the SLC6A4 gene may be related to the attack behavior, while the polymorphism of the exons in the NR2E1 gene was not detected in the polymorphism of the attack behavior. (3) in the mirror fighting test, the direct attack of the Turpan cock (Fighting) was significantly higher than the miscellaneous in the mirror fighting test. The two generation Rooster (P0.01) of the jiaozi (P0.01) was not significant in the non direct attack (Non-fighting). In the paired fight test, the direct attack (Fighting) of the Turpan chicken was significantly higher than the hybrids (P0.05), but the difference in the non direct attack (Non-fighting) behavioral variables between the two was not significant (P0.05). (4) through SLC6 The correlation between the genotypes of A4 exons and the genotypes of the exons and the matching of paired fights found that the aggression of the cockfighting and the F1 generation with the BB genotype was significantly higher than that of the other genotypes in the mirror fighting test and the paired fight test, indicating that the BB genotype of the first exon was closely related to the attack behavior. The detection of the SLC6A4 gene and NR2E1 gene expression of the weak Xinluo man laying hens and their hybrid F1 at different stages of the aggressive behavior of Turpan cockfighting was found to have a positive correlation with the attack behavior, while the gene expression of NR2E1 was negatively correlated with the attack behavior, and then the SLC6A4 and NR2E1 in Turpan were new. The SNP polymorphism of the Luo Man layer and the F1, F2 generation was analyzed, and the molecular markers associated with the attack were found in the SLC6A4 exon. The phenotype of the Turpan cockfighting and the F1 generation showed a significant difference between the attack and the phenotype of the attack line and the genotype of the first exon SLC6A4. The association analysis showed that BB genotype was correlated with aggressive behavior and could be used as a reference genotype for molecular marker selection in Turpan cockfighting.
【学位授予单位】：石河子大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S831

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