六个地方鸡品种遗传结构及其MHC基因的遗传变异研究
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摘要
我国家禽品种资源丰富,但近年来受到外来引进鸡品种的冲击,导致我国地方鸡品种群体数量日益减少。对家禽遗传资源的研究有助于了解品种资源的现状和特色,为品种资源的评价、保护和合理开发利用打下良好基础。鸡主要组织相容性复合体(major histocompatibility complex,MHC)是指16号染色体上的一组紧密连锁、高度多态的基因群编码的特异性蛋白,位于该染色体MHC B区的基因群包括B-F(Ⅰ类),B-L(Ⅱ类),B-G(Ⅳ类)三个基因座。研究表明,MHC基因多态性与家禽对多种传染病的抗性高度相关,是家禽抗病育种研究中的优选标记基因。本研究选取藏鸡、文昌鸡、固原鸡、略阳鸡、陕北鸡和太白鸡等6个地方鸡品种共584个个体作为实验材料,利用微卫星和PCR-SSCP标记技术,对6个品种的遗传结构和MHC基因在群体内的遗传变异进行分析。研究结果如下:
利用20个微卫星位点对6个地方鸡品种遗传结构进行分析,6个群体的平均等位基因数为4.9802,平均有效等位基因数为3.5464。其中,LEI0073位点等位基因最多,为12个,MCW0351和MCW0287等位基因最少,均为4个。通过哈代-温伯格平衡检验,除MCW0287、MCW0264、MCW0145和MCW0104位点外,其它位点均有群体显著偏离哈代-温伯格平衡。六个群体的平均杂合度均值为0.6717,其中藏鸡的平均杂合度最高,为0.7138,显示该品种遗传变异程度最高,遗传多样性最丰富,该品种在未来还可以进一步加大纯繁选育力度,以提高品种的遗传一致性。各群体平均多态信息含量的范围是0.5445(陕北鸡) ~0.6694(藏鸡),均大于0.5,表现为高度多态性,表明本研究所用的20个微卫星位点能够有效地应用于分析品种的遗传变异。
采用PCR-SSCP结合序列测定的方法,检测了MHC B-LBⅡ基因第二外显子在6个地方鸡群体中的遗传变异:
(1)六个品种中共检测到3种基因型,分别命名为AA型、BB型和AB型。除藏鸡群体外,AA型频率在5个鸡群体中均较高, A等位基因频率高于B等位基因频率,因此推测A等位基因为优势等位基因;Hardy-Weinberg平衡性检验结果表明所有群体均处于不平衡状态(P<0.01)。
(2)测序结果发现在176 bp的核苷酸序列中有11个变异位点,约占分析位点的6.25%,同义突变2个,其中非同义突变9个;编码的58个氨基酸中有5个氨基酸发生改变,氨基酸序列的同源性低于核苷酸序列的同源性。
(3) MHC B-LBⅡ基因在各品种内的杂合度(He)在0.1420~0.4977之间,藏鸡最高;多态信息含量(PIC)处于0.1319~0.3738之间,藏鸡最高,表明该品种的遗传资源比较丰富。
本研究结果可为我国地方鸡品种的种质特性研究提供基础性数据,为地方鸡品种资源的合理保护和利用提供科学依据。
Poultry resource is abundant in China, but it is reduced because of the introduced chicken breeds. Study on poultry resource help to understand the status and feature, provide the good base for protection of breed resources and reasonable exploition. Chicken MHC (major histocompatibility complex) is a cluster of genes located in 16 chromosome of chicken. The cluster of genes located in MHC B region includes B-F(Ⅰ), B-L(Ⅱ) and B-G(Ⅳ). Extensive studies showed that polymorphisms of MHC gene were significant correlated with the resistibility of many infectious diseases, and MHC gene is a good candidate of diseases resistance for poultry breeding.
In this study, genetic structure and variations of 584 individuals including six native breeds (Zang, Wenchang, Guyuan, Lueyang, Shanbei, and Taibei breed) were detected using microsatellite and PCR-SSCP techniques. Following is results.
1. Genetic structure and variations of six local breeds in 20 microsatellites.
1.1 The average number of alleles of 20 microsatellites in six native breeds is 4.9802, and the average number of effective alleles is 3.5464. Of all the loci, the number of alleles of LEI0073 is 12, which is maximum; both the number of alleles of MCW0351 and MCW0287 are 4, which is minimum. Except for MCW0287, MCW0264, MCW0145 and MCW0104, the Chi-Square test for goodness of fit showed Hardy-Weinberg disequilibrium is statistically significant over all the other selected loci in these populations. (p<0.05).
1.2 The average heterozygositie of populations was 0.6717. Of all the breeds, the He of Zang breed is maximum (0.7138), which indicated that both genetic variation and polymorphism are maximum. The selected pressure on this breed should be higher in later breeding practice to improve the genetic homogeneity of Zang breed.
1.3 The PIC of breeds range from 0.6694 (Zang breed) to 0.5445 (Shanbei breed), all over 0.5. The microsatellites in these breeds show higher polymorphic, which indicated the 20 microsatellites used in this study could effectively analyze genetic variations of breeds.
2. Genetic variations of exon 2 of MHC B-LBⅡgene in six native breeds.
2.1 Three genotypes (AA, AB and BB genotype) were detected in six native breeds. Except for Zang breed, Genotypic frequecy of AA is higher than other two genotyes in other five native breeds, allelic frequency of A is higher than that of B, thus allele A was dominant allele. The Chi-Square test for goodness of fit showed Hardy-Weinberg disequilibrium was statistically significant in all these populations. (p<0.01).
2.2 The sequencing results showed that there are 11 mutations in the 176 bp fragmnet, which is 6.25% in all nucleotides of this fragment. Among these mutations, 9 mutation were missense and two were synonymy. There are 5 amino acid changed in all 58 amino acid. The similarity of nucleotide sequence is lower than amino acid sequence.
2.3 The He of MHC B-LBⅡgene in six native breeds is low, ranging from 0.4977 to 0.1420. The PIC range from 0.3738 to 0.1319. The He and PIC of Zang breed was maximum, which indicates genetic resource of Zang breed is abundant.
This study could provide basic data for research on breeds characteristics of Chinese native chicken breeds and scientific basis for the protection and exploition of native chicken breeds.
利用20个微卫星位点对6个地方鸡品种遗传结构进行分析,6个群体的平均等位基因数为4.9802,平均有效等位基因数为3.5464。其中,LEI0073位点等位基因最多,为12个,MCW0351和MCW0287等位基因最少,均为4个。通过哈代-温伯格平衡检验,除MCW0287、MCW0264、MCW0145和MCW0104位点外,其它位点均有群体显著偏离哈代-温伯格平衡。六个群体的平均杂合度均值为0.6717,其中藏鸡的平均杂合度最高,为0.7138,显示该品种遗传变异程度最高,遗传多样性最丰富,该品种在未来还可以进一步加大纯繁选育力度,以提高品种的遗传一致性。各群体平均多态信息含量的范围是0.5445(陕北鸡) ~0.6694(藏鸡),均大于0.5,表现为高度多态性,表明本研究所用的20个微卫星位点能够有效地应用于分析品种的遗传变异。
采用PCR-SSCP结合序列测定的方法,检测了MHC B-LBⅡ基因第二外显子在6个地方鸡群体中的遗传变异:
(1)六个品种中共检测到3种基因型,分别命名为AA型、BB型和AB型。除藏鸡群体外,AA型频率在5个鸡群体中均较高, A等位基因频率高于B等位基因频率,因此推测A等位基因为优势等位基因;Hardy-Weinberg平衡性检验结果表明所有群体均处于不平衡状态(P<0.01)。
(2)测序结果发现在176 bp的核苷酸序列中有11个变异位点,约占分析位点的6.25%,同义突变2个,其中非同义突变9个;编码的58个氨基酸中有5个氨基酸发生改变,氨基酸序列的同源性低于核苷酸序列的同源性。
(3) MHC B-LBⅡ基因在各品种内的杂合度(He)在0.1420~0.4977之间,藏鸡最高;多态信息含量(PIC)处于0.1319~0.3738之间,藏鸡最高,表明该品种的遗传资源比较丰富。
本研究结果可为我国地方鸡品种的种质特性研究提供基础性数据,为地方鸡品种资源的合理保护和利用提供科学依据。
Poultry resource is abundant in China, but it is reduced because of the introduced chicken breeds. Study on poultry resource help to understand the status and feature, provide the good base for protection of breed resources and reasonable exploition. Chicken MHC (major histocompatibility complex) is a cluster of genes located in 16 chromosome of chicken. The cluster of genes located in MHC B region includes B-F(Ⅰ), B-L(Ⅱ) and B-G(Ⅳ). Extensive studies showed that polymorphisms of MHC gene were significant correlated with the resistibility of many infectious diseases, and MHC gene is a good candidate of diseases resistance for poultry breeding.
In this study, genetic structure and variations of 584 individuals including six native breeds (Zang, Wenchang, Guyuan, Lueyang, Shanbei, and Taibei breed) were detected using microsatellite and PCR-SSCP techniques. Following is results.
1. Genetic structure and variations of six local breeds in 20 microsatellites.
1.1 The average number of alleles of 20 microsatellites in six native breeds is 4.9802, and the average number of effective alleles is 3.5464. Of all the loci, the number of alleles of LEI0073 is 12, which is maximum; both the number of alleles of MCW0351 and MCW0287 are 4, which is minimum. Except for MCW0287, MCW0264, MCW0145 and MCW0104, the Chi-Square test for goodness of fit showed Hardy-Weinberg disequilibrium is statistically significant over all the other selected loci in these populations. (p<0.05).
1.2 The average heterozygositie of populations was 0.6717. Of all the breeds, the He of Zang breed is maximum (0.7138), which indicated that both genetic variation and polymorphism are maximum. The selected pressure on this breed should be higher in later breeding practice to improve the genetic homogeneity of Zang breed.
1.3 The PIC of breeds range from 0.6694 (Zang breed) to 0.5445 (Shanbei breed), all over 0.5. The microsatellites in these breeds show higher polymorphic, which indicated the 20 microsatellites used in this study could effectively analyze genetic variations of breeds.
2. Genetic variations of exon 2 of MHC B-LBⅡgene in six native breeds.
2.1 Three genotypes (AA, AB and BB genotype) were detected in six native breeds. Except for Zang breed, Genotypic frequecy of AA is higher than other two genotyes in other five native breeds, allelic frequency of A is higher than that of B, thus allele A was dominant allele. The Chi-Square test for goodness of fit showed Hardy-Weinberg disequilibrium was statistically significant in all these populations. (p<0.01).
2.2 The sequencing results showed that there are 11 mutations in the 176 bp fragmnet, which is 6.25% in all nucleotides of this fragment. Among these mutations, 9 mutation were missense and two were synonymy. There are 5 amino acid changed in all 58 amino acid. The similarity of nucleotide sequence is lower than amino acid sequence.
2.3 The He of MHC B-LBⅡgene in six native breeds is low, ranging from 0.4977 to 0.1420. The PIC range from 0.3738 to 0.1319. The He and PIC of Zang breed was maximum, which indicates genetic resource of Zang breed is abundant.
This study could provide basic data for research on breeds characteristics of Chinese native chicken breeds and scientific basis for the protection and exploition of native chicken breeds.
引文
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