东北民猪线粒体DNA D-loop遗传多样性与母系起源研究

为了保护东北民猪的遗传资源,对东北地区民猪群体进行线粒体DNA D-loop遗传多样性进行了调查,本研究对30头东北民猪的线粒体D-loop区部分序列(697 bp)进行扩增和测序,共检测到7个变异位点,界定了8种单倍型,核苷酸多样度(Pi)及单倍型多样度(H)分别为:0.217和0.789。结合已收录的30个亚欧猪种的线粒体D-loop序列,采用最大似然法构建的分子进化树明显分为欧洲与亚洲2个主要类群,民猪的8个单倍型都聚在亚洲分支上,并与莱芜黑猪及沂蒙黑猪较为接近,表明民猪是多母系起源群体,主要来自山东地区的华北型黑猪种群。     

中国地方猪品种的遗传多样性与聚类分析

我国拥有丰富的地方猪资源,也是世界上家猪驯化较早的中心之一。但近40年,地方猪品种受到国外引进猪种的强烈冲击,在数量和规模上均显著降低,遗传资源受到威胁。本研究利用线粒体基因组序列,对我国21个地方猪品种共计86个个体的遗传多样性进行了分析,并构建了43个地方猪品种的分子进化树。结果表明:中国地方猪线粒体D-Loop区存在序列长度多态性。86个序列共检测到32种单倍型,平均单倍型多样性(Hd)为0.966±0.006,平均核苷酸多样性(Pi)为0.00592±0.00023,遗传多样性贫乏。32种单倍型可明显的聚为2大类,但是单倍型的聚类与地理分布没有显著相关性。分子进化树显示中国地方猪与国外猪种处于2个明显不同的分支上;由于受到人类迁徙活动以及不同猪种间基因交流的影响,猪种的聚类关系与地理分布同样不存在显著相关,很难利用线粒体基因组序列对中国地方猪品种进行准确分类。     

新疆塔什库尔干牦牛mtDNA Cytb基因和D-Loop区遗传多样性及系统进化分析

为探究塔什库尔干牦牛的遗传多样性和起源进化关系,本研究通过测定和分析塔什库尔干牦牛10个个体细胞色素b基因(Cytb)和D-loop区全序列,分析多态性及构建系统进化树。结果表明:塔什库尔干牦牛Cytb基因全序列长度为1 140bp,4种核苷酸T、A、G、C的平均比例分别为27.72%、33.21%、12.92%、26.15%;在10个个体中鉴定出3种单倍型,3个变异位点,全部为转换,单倍型的多样性(Hd)为0.378,核苷酸的多样性(Pi)为0.00 205;D-loop区序列全长在890～910bp之间,4种核苷酸T、A、G、C的平均比例分别为26.12%、34.22%、25.27%、14.39%,共统计出36个SNP位点,其中转换11个,颠换21个,缺失3个,其单倍型多样性(Hd)为0.778,核苷酸多样性(Pi)为0.00 839。在基于Cytb基因的系统进化树中,塔什库尔干牦牛首先与巴州牦牛、青海牦牛聚为一类,之后与西藏牦牛、野牦牛聚为一类;在D-loop区的系统进化分析中,塔什库尔干牦牛则先与麦洼牦牛、大通牦牛聚为一类,之后与西藏牦牛、野牦牛聚为一类,展示出丰富的遗传多样性,同时进一步支持了牦牛和野牦牛划为牛亚科牦牛属的观点。     

四川黄牛品种线粒体DNA 遗传多样性研究

测定了四川黄牛5个品种的线粒体细胞色素b部分片段（420bp）42个个体以及28个个体线粒体DNA控制区（D-loop）的全序列，结合已报道的中国8个黄牛品种22个个体的线粒体DNA控制区（D-loop）全序列，分析结果表明：四川黄牛的部分Cytb基因片段有5个变异位点，7种单倍型可分为以2个单倍型为主的2大类型；线粒体DNA控制区检测到64个变异位点，28种单倍型，单倍型H1～H23为普通黄牛类型，单倍型H24～H28为瘤牛类型的单倍型。在四川黄牛中，67．86％为普通牛类型，32．14％为瘤牛类型。研究结果表明四川黄牛主要有2类母系来源。     

河北小尾寒羊mtDNA变异及遗传多样性分析

旨在分析河北小尾寒羊线粒体基因组变异和遗传多样性,采用线粒体基因组测序方法获得线粒体基因组序列,进行编码区变异分析;利用群体遗传学方法对非编码区进行遗传多样性分析。结果表明:河北小尾寒羊线粒体基因组编码区共发现183个变异位点,其中多肽编码区156个,r RNA基因19个和t RNA基因8个;多肽编码区发现同义突变128个,错义突变28个。非编码区共发现81个变异位点,核苷酸多样性为(0.022 22±0.006 93),单倍型多样性为(0.996±0.015),83只个体构成的65种单倍型,聚为3个单倍型群:A单倍型群包括35只个体,其中9只河北小尾寒羊,26只山东小尾寒羊;B单倍型群包括38只个体,其中13只河北小尾寒羊,25只山东小尾寒羊;C单倍型群为10只山东小尾寒羊,该单倍型群为一个独立类群。3个单倍型群A、B和C的频率分别为0.42、0.46和0.12。由此说明河北小尾寒羊遗传多样性较为丰富,与山东小尾寒羊存在基因交流。     

贵州3个地方猪种的mtDNA D-loop序列遗传多样性分析

为了研究3个贵州地方猪种(白洗猪、从江香猪和黔北黑猪)mt DNA D-loop区的遗传多样性,本试验采用PCR直接测序的方法测定了3个贵州地方猪种共57个个体的mt DNA D-loop区全序列。结果表明,3个贵州地方猪种mt DNA D-loop区序列长度为1254~1314bp,可分为高变区、串联重复区和保守区,串联重复片段有4种,可分为A型和B型;D-loop区的碱基含量特点为A+T明显高于G+C;在白洗猪、从江香猪和黔北黑猪的高变区分别检测到6、6、8个均为转换的突变位点,分别定义了4、4、6种单倍型,单倍型多样度(Hd)为0.6386~0.8974,核苷酸多样度为0.0025~0.0035,3个贵州地方猪种均表现出较为丰富的遗传多样性。     

淮南猪线粒体DNA D-loop遗传多样性及母系起源研究

为探究淮南猪的遗传多样性和母系起源,本实验对32头淮南猪线粒体DNA D-loop区序列进行PCR扩增和测序,并结合GenBank中国内外猪种及黄河流域古家猪D-loop序列,分析淮南猪线粒体多态性及其起源。结果显示:淮南猪1 170 bp的D-loop序列核苷酸多样性(Pi)及单倍型多样性(Hd)分别为0.004 25和0.968;32个测序结果共检测到27个多态位点,界定了20种单倍型;中性检验表明淮南猪在驯化进程中曾发生过种群扩张;邻接法(NJ)构建分子系统发育树明显将群体分为国内和国外2个类群,淮南猪20个单倍型都聚集在国内分支上;进一步Network分析显示,淮南猪单倍型在国内猪种中呈广泛分布,但与华中、华北型猪种基因交流较多,且与黄河流域古家猪遗传关系较近,表明淮南猪为多母系起源,主要来自于黄河流域古家猪。     

西藏小型猪线粒体DNA控制区的分子遗传学研究

通过扩增102头西藏小型猪以及16头巴马小型猪、17头贵州香猪的线粒体DNA控制区,测序并与国内其他猪种进行比较,研究西藏小型猪的遗传标记以及与其他国内地方猪种的亲缘关系.结果显示,两藏小型猪线粒体DNAD-loop区分3个区域.串联重复序列区处于中间位置,包含有15～29个10 bp的重复片段,分为A、B 2种类型.3'端340 bp,与国内其他猪种的序列相同,比较保守;D-loop 5'端704 bp,共有22个变异位点.由22个变异位点中归纳出25个单倍型,其中有2种主要的单倍型,分别占34.4%和36.6%.根据3个转换位点:305、500、691,将西藏小型猪分成了2组,几乎与串联重复序列所分的A、B 2组类型相对应.与西藏小型猪相比,巴马小型猪和贵州香猪D-loop 5'端变异位点较少,分别只有4种和2种单倍型,串联重复区也只有1个类型.说明西藏小型猪可能有2个母系祖先,并且与我国西南地区的猪种有较近的亲缘关系;不同的串联重复片段类型和5'端的变异位点可以联合组建西藏小型猪的遗传标记.     

苏姜猪mtDNA D-loop序列的遗传多样性分析

【目的】通过对苏姜猪线粒体DNA(mtDNA) D-loop高变区Ⅰ的序列扩增测序,探究苏姜猪种群的种质资源特性和遗传多样性。【方法】采集苏姜猪核心群100头经产母猪耳组织,提取DNA后扩增苏姜猪核心群母猪mtDNA D-loop高变区Ⅰ的基因片段并测序,运用NCBI在线BLAST对测序序列与参考序列进行比对、校正并寻找同源序列,确定mtDNA D-loop高变区Ⅰ序列的长度和位置;使用DnaSP v.5.10.1软件统计获得群体中核苷酸多态位点和变异位点数量,分析单倍型多样度(Hd)、核苷酸多样性(Pi)、平均核苷酸差异度(K)等参数;使用Mega 7.0软件对不同单倍型进行基于Kimura’s 2-prarmetermodel的遗传距离计算,采用邻接法(Neighbor-Joining, NJ)对苏姜猪的4个单倍型、17个中国地方猪种、欧洲家猪(登录号：AF034253.1)的mtDNA D-loop高变区Ⅰ序列构建系统发育树。【结果】对100头苏姜猪的mtDNA D-loop序列高变区Ⅰ进行扩增测序,获得长度为429 bp的有效序列,获得的序列中,AT含量为63.1%,GC含量为...     

重庆本地山羊资源——铜梁麻羊的分子系统学分析

为了分析铜梁麻羊线粒体DNA(mtDNA)D-loop区遗传多样性,鉴别其是否为一个新的山羊遗传资源,试验采用生物信息学方法,对8只铜梁麻羊线粒体D-loop区序列和川渝13个山羊品种的线粒体D-loop区共88条序列一起进行了分子系统学分析。结果表明:铜梁麻羊线粒体D-loop区序列为1 212 bp,共发现39个变异位点,共定义出4种品种特有单倍型,其核苷酸多样度为0.015±0.010,单倍型多样度为0.700±0.008;品种间的遗传距离显示,铜梁麻羊与成都麻羊、黔北麻羊的遗传距离较大;聚类分析显示,铜梁麻羊与成都麻羊和黔北麻羊同在一个支系,但亲缘关系较远。说明铜梁麻羊在外貌和基因层次上均与现有的麻羊品种有明显的区别,很可能是一个新的山羊类群,值得进一步保护和开发。     

Genetic Diversity and Phylogenetic Relationships of the Complete Mitochondrial Genome in Chinese Indigenous Pig Breeds

In order to explore the genetic diversity and the classification status of the Chinese domestic pigs at the molecular level, the complete mitochondrial DNA (mtDNA) genomic sequences of 22 Chinese indigenous pig breeds from 6 types were downloaded from GenBank for the analysis using the bioinformatics method. The polymorphism of nucleic acid sequences was analyzed,the molecular phylogenetic tree based on D-loop sequences, Cytb gene, complete coding region mtDNA sequences and the Median-Joining network of the mtDNA D-loop haplotypes were constructed for 22 pig breeds from 6 types. The results showed that a total of 144 mutation sites among 22 pig breeds were detected, 22 haplotypes were discovered, which indicated that Chinese indigenous pig breeds had abundant genetic diversity. According to the complete mtDNA genomic sequences analysis, which suggested that the main mutations was transition, the transition/transversion ratio was greater than 2.0 and all mutations were in line with the neutral mutation. Our findings clearly demonstrated that there was a near genetic distance among 6 types, meanwhile, the shared haplotypes were found. The phylogenetic tree showed that Chinese indigenous pigs from 6 types were diverged from two ancestors.The results indicated that mtDNA D-loop and Cytb gene might serve as molecular markers for phylogenesis, origin and evolution.     

The phylogenetic status of typical Chinese native pigs: analyzed by Asian and European pig mitochondrial genome sequences

China is one of the most diverse countries, which have developed 88 indigenous pig breeds. Several studies showed that pigs were independently domesticated in multiple regions of the world. The purpose of this study was to investigate the origin and evolution of Chinese pigs using complete mitochondrial genomic sequences (mtDNA) from Asian and European domestic pigs and wild boars. Thirty primer pairs were designed to determine the mtDNA sequences of Xiang pig, Large White, Lantang, Jinhua and Pietrain. The phylogenetic status of Chinese native pigs was investigated by comparing the mtDNA sequences of complete coding regions and D-loop regions respectively amongst Asian breeds, European breeds and wild boars. The analyzed results by two cluster methods contributed to the same conclusion that all pigs were classified into two major groups, European clade and Asian clade. It revealed that Chinese pigs were only recently diverged from each other and distinctly different from European pigs. Berkshire was clustered with Asian pigs and Chinese pigs were involved in the development of Berkshire breeding. The Malaysian wild boar had distant genetic relationship with European and Asian pigs. Jinhua and Lanyu pigs had more nucleotide diversity with Chinese pigs although they all belonged to the Asian major clade. Chinese domestic pigs were clustered with wild boars in Yangtze River region and South China.     

陆川猪mtDNA D-loop序列遗传多样性分析

本研究测定了陆川猪17个个体mtDNA D-loop高变区序列,共检测到5个单倍型,单倍型多样度和核苷酸多样度分别为0.640和0.0074,表明陆川猪有较高的遗传多样性。结合已报道的中国地方猪种的mtDNA D-loop序列进行分析,结果发现,陆川猪和属于华中型的宁乡猪、大花白猪有很近的遗传关系,但与传统上同属华南型的香猪和滇南小耳猪的遗传关系较远,这与传统的分类方法不一致。     

The Genetic Diversity and Phylogenetic Relationships of Five Guangxi Pig Breeds and Three Exotic Pig Breeds by Analysis of mtDNA D-loop Sequences

In order to compare the genetic diversity and phylogenetic relationships between Guangxi pig breeds and three exotic pig breeds,one pair of primers was used to amplify 85 mtDNA D-loop sequences of five Guangxi pig breeds (Bama Xiang pig,Dongshan pig,Guizhong spotted pig,Luchuan pig,Huanjiang Xiang pig),and we downloaded 45 mtDNA D-loop sequences of three exotic pig breeds (Duroc,Yorkshire,Landrace) from GenBank.Then the haplotype,genetic diversity and phylogenetics analysis were carried out.The results showed that 38 haplotypes were identified in 130 sequences,and the mean distances among Guangxi pig breeds were smaller than that among Duroc,Landrace and Guangxi pig breeds.The NJ phylogenetic tree classified all samples into two clusters.One contained all of Guangxi pig breeds and partial Yorkshire,the other was composed wholly of exotic pig breeds,which mean there were two distinct maternal origins.In the cluster of Guangxi pig breeds,Luchuan pig were clustered,and the others were rather dispersive.It indicated that Luchuan pig had simple maternal bloods,and the other breeds had complex maternal bloods,and they had gene flow with each other.The maternal pig resources in Guangxi were not affected by the introduction of exotic pig breeds,and they were still suitable for use as a female parent in their development and utilization.     

几个地方绵羊品种线粒体DNA(mtDNA)细胞色素b基因多态性研究

用12种限制性内切酶分析了6个地方绵羊品种线粒体DNA（mtDNA）细胞色素b（cytb）基因的RFLP。结果表明，在71只绵羊个体中，检出的14种限制性态型可归结成4种单倍型，其间的差异主要来源于几个限制性位点的点突变；4种单倍型间和6个绵羊群体间的平均遗传距离（D）分别为0．412％和0．041％，遗传多态程度（π）为0．028％，说明遗传多样性较为贫乏；单倍型聚类结果和群体聚类结果提示，我国地方绵羊品种的起源可能是单一的。     

深县猪mtDNA CytB基因的遗传多样性与母系起源研究

为探讨深县猪群体内的遗传多样性及母系起源分化,采用PCR直接测序法测定了40头深县猪的mtDNA CytB基因长1 140 bp片段的全序列,并结合GenBank已公布的15个猪种mtDNA CytB基因全序列,采用邻接法构建深县猪、部分其他地方猪种和引进猪种的系统发育树。结果显示,在1 140 bp长的序列中A、T、G、C的含量分别为25.93%、31.72%、28.90%及13.45%,其中A+T的含量(57.65%)高于G+C的含量(42.35%);共发现2个变异位点,无插入和缺失突变,全部为转换位点,其中简约信息位点2个,未发现单一信息位点。40条序列共定义了3种单倍型,单倍型多样性(HD)为0.312,核苷酸多样性(Pi)为0.00026,平均核苷酸差异数(K)为0.327。在深县猪mtDNA CytB基因全序列NJ进化树中,三种单倍型聚为同一分支,母系来源单一,与莱芜猪、大蒲莲猪等山东品种遗传距离较近。结果表明,深县猪群体内遗传多样性比较贫乏,与山东地区的华北型黑猪种群有更近的亲缘关系。     

五指山猪线粒体控制区序列分析及遗传多样性研究

为了研究五指山猪遗传多样性和系统地位,本研究采用PCR产物直接测序法得到55条五指山猪线粒体控制区(mtDNA D-Loop)全序列,并结合GenBank中发表的1条五指山猪mtDNA D-Loop全序列进行结构分析、遗传多样性研究及系统发育分析。结果发现,56条序列中出现21次保守区变异,即"TTATAAAACAC"序列重复,共定义13个单倍型,发现14个变异位点,单倍型多样度(Hd)和核苷酸多样度(Pi)分别为0.838和0.00334,表明五指山猪遗传多样性较丰富。五指山猪线粒体控制区序列构建的系统发育树和Network网络分析呈现两大分支,推测有两个母系起源,且与GenBank中其他23个猪种的聚类结果表明,五指山猪与长江流域以南地区的猪种有着较近的亲缘关系。     

Complete Mitochondrial DNA D-Loop Sequence and Genetic Diversity of Wuzhishan Pig

In order to research the genetic diversity and phylogenies of Wuzhishan pig, 56 complete mtDNA D-Loop were analyzed.55 sequences were amplified by polymerase chain reaction (PCR) and 1 sequence was downloaded from GenBank.The results showed that repetitive sequence "TTATAAAACAC" appeared in the conservative regions of 21 mtDNA D-Loop sequences.13 haplotypes and 14 variable sites were observed in 56 sequences.Haplotype diversity and nucleotide diversity (Hd =0.838 and Pi=0.00334) indicated that Wuzhishan pig had high genetic diversity.Two branches in the phylogenetic tree and Network analysis diagram indicated that there were two maternal origins in Wuzhishan pig.The phylogenetic tree of Wuzhishan pig and other 23 species showed that Wuzhishan pig had close genetic relationship with the pig breeds in the south region of the Yangtze river.     

建昌马mtDNA D-loop区遗传多样性及系统进化分析

试验旨在以线粒体DNA(mitochondrial DNA,mtDNA)为切入点,研究建昌马的母系遗传多样性与系统进化。从建昌马(n=39)血液中提取基因组DNA,用PCR方法扩增mtDNA D-loop区并直接测序,分析其高变区247 bp序列信息,统计mtDNA D-loop区的单倍型及变异位点,计算单倍型多样性(haplotype diversity,Hd)、核苷酸多样性(nucleotide diversity,Pi)和平均核苷酸变异数(average number of nucleotide differences,K)。构建包括建昌马在内的19个品种马的NJ系统进化树,计算各品种间的遗传距离。结果显示,试验获得了清晰的PCR扩增产物,并通过直接测序方法获得了约1200 bp的序列。39匹建昌马mtDNA D-loop区247 bp序列(其中1个样品缺失1 bp)的AT碱基含量为61.45%,属AT碱基对富集区,检测到33个多态性位点,共显示26种单倍型,其中4种为共享单倍型,且Hap7和Hap1为优势单倍型,单倍型多样性为0.947,核苷酸多样性为0.02399,平均核苷酸变异数为5.901,显示丰富的母系遗传多样性;NJ系统进化树显示,建昌马分布在A、C、D、E、F、G共6个支系中,约50%的样品分布在A支系,显示出复杂的母系起源;建昌马与关中马的遗传距离最小(0.021),其次是三河马、文山马、韩国车巨马(0.024),与韩国济州岛马遗传距离最大(0.032)。本研究结果表明,建昌马的mtDNA D-loop高变区遗传多样性丰富,具有多个母系起源,且A支系占有明显优势,与关中马、文山马可能有共同的母系起源。     

江淮水牛线粒体D-loop区和Cytb基因遗传多态性及系统发育分析

为研究安徽江淮水牛群体的分子种质特性,本试验测定了江淮水牛2个亚群共82个个体的线粒体D-loop区和细胞色素b基因完整序列,分析了序列遗传多态性及系统进化关系,并结合GenBank中已发表的141条中国水牛D-loop序列,进行了联合分析。结果在D-loop区内共发现核苷酸多态位点91个,组成104个单倍型,其中32个是在江淮水牛中新发现的单倍型。总体mtDNA D-loop区核苷酸多样性为(0.015 43±0.001 42),单倍型多样性为(0.948±0.009)。其中,江淮水牛的mtDNA D-loop区核苷酸多样性为(0.014 89±0.002 32),单倍型多样性为(0.955±0.013),表明其群体遗传多样性丰富,群体变异性水平与中国其他水牛群体接近。根据线粒体D-loop区单倍型构建系统树和进化网络,显示江淮水牛存在沼泽型水牛线粒体支系A和B,表明其具有2个线粒体母系来源,其中B支又分为b1和b2两个亚支;针对细胞色素b基因的进化分析也支持这一结论。这些研究结果为今后开展江淮水牛遗传资源的保护和利用提供了客观依据。