Hypervariability of the D-loop region in mitochondrial DNA of Russian sturgeon <Emphasis Type="Italic">Acipenser gueldenstaedtii</Emphasis> (Acipenseriformes,Acipenseridae)

The structure of the D-loop region in mitochondrial DNA (mtDNA) of Russian sturgeon Acipenser gueldenstaedtii from the Azov Sea population was studied with the method of direct sequencing. Interindividual heteroplasmy of the length of mtDNA in the region of D-loop realized by the presence of a different number of tandem repeats (82 pairs of bases) was found. Analysis of tandem repeats in the D-loop region in mtDNA in the studied sample (28 individuals) revealed eight mitotypes differed in the pattern of nucleotide substitution and in the number of tandem repeats (2, 3, and 4 repeats). Revealed mitotypes can be considered as potential genetic markers for different biological groups, schools, or seasonal races of A. gueldenstaedtii.     

4种鲟鱼养殖亲鱼群体遗传多样性分析

近年鲟鱼人工繁殖技术促进了鲟鱼养殖业的发展,但在选育初期未对后备亲鱼进行遗传背景分析,为了抑制种质资源退化和防止近交衰退,繁育场需要对现有后备亲鱼进行遗传背景分析.本文利用线粒体控制区(D-loop)部分序列对4种养殖鲟鱼后备亲鱼群体(共120个样本)的遗传多样性进行了分析.研究发现不同种鲟鱼D-loop部分序列长度不...     

Mitochondrial DNA Length Variation and Heteroplasmy in Populations of White Sturgeon (Acipenser Transmontanus)

Southern blot analysis was used to quantify the extent of mtDNA D-loop length variation in two populations of white sturgeon, Acipenser transmontanus. Over 42% of individuals were heteroplasmic for up to six different mtDNA length variants attributable to varying copy numbers of an 82-bp repeat sequence. Chi-square analyses revealed that the frequencies of length genotypes and the incidence of heteroplasmy were significantly different between Fraser and Columbia River sturgeon populations but not between restriction site haplotypes. Heteroplasmic fish have, on average, higher copy number than homoplasmic fish. Forty-five of 101 homoplasmic individuals carry only a single copy of the repeat, while none of the 73 heteroplasmic fish has the single repeat as the predominant variant. On the basis of differences in frequency distributions of copy number within and between fish, we suggest that (1) heteroplasmy is maintained by high recurrent mutation of multiple copy genomes, favoring increased copy number and (2) the mutation pressure toward higher copy number heteroplasmy is partially offset by selection to reduced genome size and segregation to the homoplasmic condition.     

Intraspecific DNA sequence variation of the mitochondrial control region of white sturgeon (Acipenser transmontanus)

Intraspecific sequence variation in the D-loop region of mtDNA in white sturgeon (Acipenser transmontanus), a relict North American fish species, was examined in 27 individuals from populations of the Columbia and Fraser rivers. Thirty-three varied nucleotide positions were present in a 462-nucleotide D-loop sequence, amplified using the polymerase chain reaction. Bootstrapped neighbor-joining and maximum- parsimony trees of sequences from 19 haplotypes suggest that the two populations have recently diverged. This is consistent with the hypothesis that the Columbia River, a Pleistocene refugium habitat, was the source of founders for the Fraser River after the last glacial recession. On the basis of a divergence time of 10-12 thousand years ago, the estimated substitution rate of the white sturgeon D-loop region is 1.1-1.3 x 10(-7) nucleotides/site/year, which is comparable to rates for hypervariable sequences in the human D-loop region. Furthermore, the ratio of mean percent nucleotide differences in the D- loop (2.27%) to that in whole mtDNA (0.54%, as estimated from restriction-enzyme data) is 4.3, which is similar to the fourfold-to- fivefold-higher substitution rate estimated for the human D-loop. The high nucleotide substitution rate of the hypervariable region indicates that the vertebrate D-loop has potential as a genetic marker in molecular population studies.      

Heteroplasmy of Short Tandem Repeats in Mitochondrial DNA of Atlantic Cod, Gadus Morhua

The mitochondrial DNA of the Atlantic cod (Gadus morhua) contains a tandem array of 40-bp repeats in the D-loop region of the molecule. Variation among molecules in the copy number of these repeats results in mtDNA length variation and heteroplasmy (the presence of more than one form of mtDNA in an individual). In a sample of fish collected from different localities around Iceland and off George's Bank, each individual was heteroplasmic for two or more mtDNAs ranging in repeat copy number from two (common) to six (rare). An earlier report on mtDNA heteroplasmy in sturgeon (Acipenser transmontanus) presented a competitive displacement model for length mutations in mtDNAs containing tandem arrays and the cod data deviate from this model. Depending on the nature of putative secondary structures and the location of D-loop strand termination, additional mechanisms of length mutation may be needed to explain the range of mtDNA length variants maintained in these populations. The balance between genetic drift and mutation in maintaining this length polymorphism is estimated through a hierarchical analysis of diversity of mtDNA length variation in the Iceland samples. Eighty percent of the diversity lies within individuals, 8% among individuals and 12% among localities. An estimate of theta = 2N(eo) mu greater than 1 indicates that this system is characterized by a high mutation rate and is governed primarily by deterministic dynamics. The sequences of repeat arrays from fish collected in Norway, Iceland and George's Bank show no nucleotide variation suggesting that there is very little substructuring to the North Atlantic cod population.     

Sequence Heteroplasmy of D-Loop and rRNA Coding Regions in Mitochondrial DNA from Holstein Cows of Independent Maternal Lineages

A mitochondrial DNA (mtDNA) fragment containing the D-loop, phenylalanine tRNA, valine tRNA, and 12S and 16 rRNA genes was cloned and sequenced from 36 cows of 18 maternal lineages to identify the polymorphic sites within those regions and to detect the existence of heteroplasmic mtDNA in cows. Seventeen variable sites were observed within the D-loop and rRNA coding regions of bovine mtDNA within a 2.5-kb span. The hypervariable sites in the D-loop and rRNA coding regions were identified at nucleotide positions 169, 216, and 1594. Heteroplasmic mtDNA (variable mtDNA within a tissue) existed extensively in cows and was detected within the above regions in 11 of 36 cows sequenced. The insertion, deletion, and nucleotide transversion polymorphisms were found only in homopolymer regions. Heteroplasmy was observed frequently and seemingly is persistent in cattle. Though heteroplasmy was demonstrated, most lineages and mtDNA sites showed no heteroplasmy.     

MtDNA substitution rate and segregation of heteroplasmy in coding and noncoding regions

The mitochondrial DNA (mtDNA) substitution rate and segregation of heteroplasmy were studied for the non-coding control region (D-loop) and 500 bp of the coding region between nucleotide positions 5550 and 6050, by sequence analysis of blood samples from 194 individuals, representing 33 maternal lineages. No homoplasmic nucleotide substitutions were detected in a total of 292 transmissions. The estimated substitution rate per nucleotide per million years for the control region (micro>0.21, 95% CI 0-0.6) was not significantly different from that for the coding region (micro>0.54, 95% CI 0-1.0). Variation in the length of homopolymeric C streches was observed at three sites in the control region (positions 65, 309 and 16,189), all of which were in the heteroplasmic state. Segregation of heteroplasmic genotypes between generations was observed in several maternal pedigrees. At position 309, a longer poly C tract length was strongly associated with a higher probability for heteroplasmy and rapid segregation between generations. The length heteroplasmy at positions 65 and 16,189 was found at low frequency and was confined to a few families.     

The Genetic Diversity of mtDNA D-loop and the Origin of Chinese Goats

The complete sequences of mitochondrial DNA D-loop of 128 individuals in nine Chinese goat (Capra hircu) breeds were analyzed by DNA sequencing technology. The results show that the length of mtDNA D-loop in Chinese goats is 1 212-1 213 bp. There are 102 polymorphic sites, accounting for 8.42% of 1 212 bp sequence. Ninety-two mtDNA haplotypes were determined. The haplotype diversity and nucleotide diversity are 0.9333-1 .0000 and 0.7062%-1.8265%, respectively. The results indicate that the genetic diversity of Chinese goats is very abundant. The NJ tree indicates that Chinese goats have two types of maternal origins from lineage A and lineage B. The possibility of lineage B originating from China is also discussed.     

中国山羊mtDNA D-loop遗传多样性及其起源研究

采用DNA测序技术分析了中国9个山羊品种（板角山羊、成都麻羊、贵州黑山羊、贵州白山羊、黔北麻羊、马头山羊、陕南白山羊、黄淮山羊和雷州山羊）共计128个个体的mtDNA D-loop全序列。结果表明：山羊mtDNA D-loop全序列长度为1212-1213bp,检测到102个变异位点,约占分析位点总数的8．42％,可变位点中转换占99个,颠换2个,1个转换/颠换共存;界定了92种单倍型,有78种为各品种独享单倍型,另外14种为群体内或群体间共享单倍型。9个山羊品种单倍型多样度为0．9333-1．0000,核苷酸多样度为0．7062％-1．8265％,表明中国山羊品种遗传多样性丰富。根据92种mtDNA单倍型构建了中国山羊的NJ分子系统树,聚类表明,中国山羊mtDNA D-loop序列单倍型分为支系A和支系B两大类。支系A包括75种单倍型,代表95个样本,占总数的74．22％;支系B包括17种单倍型,代表33个样本,占总数的25．78％,说明中国山羊存在支系A和支系B两大母系起源。对中国山羊mtDNA D-loop的支系A和支系B进行核苷酸不配对分布曲线分析和Fu的Fs中性检验,分析表明,支系A的分布曲线呈单峰形,Fs值为-24．6491,P值为0．0000,显著偏离中性,表明山羊支系A曾经历群体扩张;支系B呈近似双峰分布,Fs值为-3．3947,P值为0．0980,中性检验差异不显著,表明山羊支系B没有经历群体扩张,群体大小保持相对稳定。山羊支系B可能起源于中国。     

Population genetics of shortnose sturgeon Acipenser brevirostrum based on mitochondrial DNA control region sequences

Shortnose sturgeon is an anadromous North American acipenserid that since 1973 has been designated as federally endangered in US waters. Historically, shortnose sturgeon occurred in as many as 19 rivers from the St. John River, NB, to the St. Johns River, FL, and these populations ranged in census size from 10(1) to 10(4), but little is known of their population structure or levels of gene flow. We used the polymerase chain reaction (PCR) and direct sequence analysis of a 440 bp portion of the mitochondrial DNA (mtDNA) control region to address these issues and to compare haplotype diversity with population size. Twenty-nine mtDNA nucleotide-substitution haplotypes were revealed among 275 specimens from 11 rivers and estuaries. Additionally, mtDNA length variation (6 haplotypes) and heteroplasmy (2-5 haplotypes for some individuals) were found. Significant genetic differentiation (P < 0.05) of mtDNA nucleotide-substitution haplotypes and length-variant haplotypes was observed among populations from all rivers and estuaries surveyed with the exception of the Delaware River and Chesapeake Bay collections. Significant haplotype differentiation was even observed between samples from two rivers (Kennebec and Androscoggin) within the Kennebec River drainage. The absence of haplotype frequency differences between samples from the Delaware River and Chesapeake Bay reflects a probable current absence of spawning within the Chesapeake Bay system and immigration of fish from the adjoining Delaware River. Haplotypic diversity indices ranged between 0.817 and 0.641; no relationship (P > 0.05) was found between haplotype diversity and census size. Gene flow estimates among populations were often low (< 2.0), but were generally higher at the latitudinal extremes of their distribution. A moderate level of haplotype diversity and a high percentage (37.9%) of haplotypes unique to the northern, once-glaciated region suggests that northern populations survived the Pleistocene in a northern refugium. Analysis of molecular variance best supported a five-region hierarchical grouping of populations, but our results indicate that in almost all cases populations of shortnose sturgeon should be managed as separate units.     

Post-bottleneck mtDNA diversity in a free-living population of European bison: implications for conservation

A free-living population of European bison Bison bonasus in the Białowieża Primeval Forest originated from only seven founder animals after a severe bottleneck that occurred at the beginning of the 20th century. Consequently, the contemporary population of the species is characterized by low genetic diversity. We studied a total of 195 individuals (127 males and 68 females). A 1429 bp fragment of mitochondrial DNA (mtDNA) including the D-loop region was analyzed in 87 individuals and revealed only three distinct haplotypes. Nucleotide ( π ) and haplotype ( H _d) diversity values were estimated for the European bison and were compared with π and H _d estimated from three individuals of American bison Bison bison . Very low diversity values were found in the European bison in comparison with the diversity values found in the American bison. The low mtDNA variability in the European bison is in concordance with theoretical expectations for a species that has undergone a severe and recent bottleneck. A management strategy for the preservation of the rare and very rare haplotypes present in the Białowieża population of the European bison is discussed. Furthermore, all 195 individuals were investigated for heteroplasmy involving these three haplotypes, in order to detect a possible association between heteroplasmy and the incidence of males affected by posthitis , a disease that affects the male reproductive organs, leading to necrotic lesions. Heteroplasmy was found in 15 females, in 17 males affected by posthitis and in 11 non-affected males, and no significant association was found.     

多浪羊mtDNA D-loop区遗传多样性及系统发育分析

以多浪羊为研究对象,分析绵羊线粒体D-loop区的遗传多样性,为研究多浪羊的起源和进化历史奠定基础。结果显示,多浪羊线粒体DNA D-loop序列长度为945~1 039 bp,A、T、G和C含量分别为29.4%、27.7%、17.7%和25.1%,其中A+T为57.1%,G+C为42.8%。研究获得了26种单倍型,56个多态位点,其中单一多态位点42个,简约信息位点14个。平均核苷酸差异数k为5.289,核苷酸多样度Pi为0.02 415,核苷酸多样度较低,说明多浪羊的遗传多样性贫乏,应采取重点措施予以保护。另外研究发现多浪羊经历过群体扩张,其母系起源除A、B和C世系外,可能存在D或E世系。     

Phylogeography of sable (Martes zibellina L. 1758) in the southeast portion of its range based on mitochondrial DNA variation: highlighting the evolutionary history of the sable

To reveal phylogeographic features of sable (Martes zibellina) in the southeast part of its range, we analyzed variability of the mitochondrial DNA (mtDNA) cytochrome b gene, tRNA (Pro), tRNA (Thr) and control region (D-loop) sequences from 78 specimens in populations of the Russian Far East, northeast China, and Mongolia. Our results revealed the presence of 49 different haplotypes split into two major phylogenetic groups—clades A and B, the latter separated into two clades, B1 and B2. Comparative analysis of D-loop haplotypes in populations originating from the southeast (Russian Far East, China and Mongolia) and the west (northern Urals) portions of sable range indicated that all three mtDNA clades were present in different regional groups. However, highest diversity of clade B1 in northeast China and its nearly complete absence from the Urals suggest that the southeast sable range, being a refuge during Pleistocene glacial periods, can be considered the center of genetic diversification and possibly origin of this species. All divergence estimates fall within the Pleistocene suggesting that Quarternary glaciations played an important role in phylogeographic differentiation of sable.     

贵州黄牛mtDNA D-loop 遗传多样性研究

对贵州4个地方黄牛品种共计82个个体的线粒体DNA D-loop区全序列910 bp进行分析,检测到31种单倍型,其核苷酸多态位点65个,约占所测核苷酸总长的7.14%,其中有62个转换,2个颠换,1个转换/颠换共存。贵州4个黄牛品种mtDNA D-loop区核苷酸多样度（π值）为2.16%～2.61%,单倍型多样度（H）为0.695～0.909,表明贵州黄牛mtDNA遗传多样性比较丰富。根据单倍型构建了贵州4个黄牛品种的NJ分子系统树。聚类表明,贵州黄牛有普通牛和瘤牛2大母系起源,其影响较为均一。并探讨了用核苷酸多样度π值的大小来衡量黄牛群体遗传分化程度的可行性。      

Bovine mtDNA D-loop haplotypes exceed mutations in number despite reduced recombination: an effective alternative for identity control

Mitochondrial (mt) DNA D-loop heterogeneity, haplotype distribution and possible sub-population structures within the relevant populations are important for DNA-based traceability. To gain insight into this distribution, we compared 1515 Bos taurus mtDNA D-loop sequences available from GenBank to 219 sequences that we sequenced de novo. A pronounced ambiguous trace typical of C-track length heteroplasmy was encountered in 5% of the samples, which were excluded from the analysis. Previously undescribed mutations and haplotypes were observed in 6% and 63% of the sequences, respectively. B. taurus haplotypes divided into the taurus, indicus and grunniens types and 302 variable sites formed the 858 taurus haplotypes detected. Fifty-five sites displayed a complex level of variation. As each level represents an independent mutation event, a total of 399 mutations were traced, which could potentially explain independent formation of less than half (47%) of the haplotypes encountered: most haplotypes were derived from different combinations of these mutations. We suggest that a mutational hotspot may explain these results and discuss the usefulness of mtDNA for identity and maternity assurance.     

小口白甲鱼都柳江种群mtDNA D环的序列变异及遗传多样性

采用PCR结合DNA测序技术,测定分析了易危鱼类小口白甲鱼(Onychostoma lini)都柳江种群36个个体mtDNA D环约470bp序列的变异及遗传多样性。结果表明,在36个个体中,该序列的长度为469～475bp,其碱基组成为A+T的平均含量(68.4%)高于G+C(31.6%)。共检测到25个多态位点,其中转换19个、颠换6个。核苷酸多样性(π)为0.00575,平均核苷酸差异数(K)为2.695。36个个体分属5个单倍型,单倍型多样度(Hd)为0.260,单倍型间的平均遗传距离(P)为0.026。5个单倍型构建的UPGMA系统树聚为2个分支。目前小口白甲鱼都柳江种群mtDNA D环序列存在着较丰富的变异和遗传多样性。     

African DNA Lineages in the Mitochondrial Gene Pool of Europeans

Mitochondrial DNA (mtDNA) nucleotide sequences of African origin are found in various European populations at a low frequency (on average, less than 1%). Data on mtDNA variation in Eurasian and African populations have been analyzed, and African mtDNA lineages have been found in Europeans. It has been demonstrated that, despite the high diversity of mtDNA haplotypes of African origin in Europeans, few monophyletic clusters of African lineages are characterized by long-term diversity formed in Europe. Only two such mtDNA clusters (from haplogroups L1b and L3b) have been found, their evolutionary age not exceeding 6500 years. European and African populations have been compared with respect to the frequency distributions of the alleles of autosomal microsatellite loci found in Russian carriers of African mtDNA haplotypes. It has been demonstrated that alleles typical of Europeans are characteristic of the autosomal genotypes of these Russian individuals.     

Analysis of heteroplasmy in the major noncoding region of mitochondrial DNA in the blood and atherosclerotic plaques of carotid arteries

For identification of somatic mitochondrial DNA (mtDNA) mutations, the mtDNA major noncoding region (D-loop) sequence in blood samples and carotid atherosclerosis plaques from patients with atherosclerosis was analyzed. Five point heteroplasmic positions were observed in 4 of 23 individuals (17%). Only in two cases could heteroplasmy have resulted from somatic mutation, whereas three heteroplasmic positions were found in both vascular tissue and blood. In addition, length heteroplasmy in a polycytosine stretches was registered at nucleotide positions 303–315 in 16 individuals, and also in the 16184–16193 region in four patients. The results suggest that somatic mtDNA mutations can occur during atherosclerosis, but some heteroplasmic mutations may appear in all tissues, possibly being inherited.     

中国黄牛mtDNA D-loop遗传多样性及起源

黄牛自古以来就是我国一个重要的畜种,其经济、文化价值很高。我国是世界上黄牛品种资源最丰富的国家之一。据《中国牛品种志》介绍,把一些地区同种异名的黄牛品种合并以后,尚有28个地方黄牛品种,按照其地理分布区域分为北方黄牛、中原黄牛和南方黄牛三大类型(邱怀,1986)。如果把中国地方黄牛品种分得更细,则有49个固有品种(常洪,1995)。关于中国黄牛的起源,历来有不同的观点。一般认为,中国黄牛是多元起源的,但究竟起源于哪几个牛种,观点不一(陈宏等,1993;于汝梁等,1993;Yu et al.,1999;陈幼春,1990)。主要的观点有:(1)中国黄牛主要起源于…     

Polymorphism of the mitochondrial DNA control region in eight sturgeon species and development of a system for DNA-based species identification

Intraspecific and interspecific nucleotide sequence variations of the mtDNA control region (D-loop) were studied with mtDNAs isolated from tissue specimens of more than 1400 sturgeons of nine species: Russian sturgeon Acipenser gueldenstaedtii, Persian sturgeon A. persicus, Siberian sturgeon A. baerii, Amur sturgeon A. schrenkii, Fringebarbel sturgeon A. nudiventris, sterlet A. ruthenus, stellate sturgeon A. stellatus, beluga Huso huso, and kaluga H. dauricus. The results were used to analyze the interspecific variation of the mtDNA control region in the given set of species and to develop a test system of ten species-specific primers, which allowed species identification from noninvasive tissue samples, spawn, and food products of eight species. The system proved suitable for multiplex PCR. A method was developed for the first time to reliably differentiate the A. baerii mitotype and the baerii-like mitotype of A. gueldenstaedtii. It was found that, although genetically separate, A. gueldenstaedtii and A. persicus are relatively young species and have common mitochondrial haplotypes, precluding their identification via mtDNA analysis alone. To develop a system for species identification of A. gueldenstaedtii and A. persicus, it is necessary to study the polymorphism of nuclear markers.