Phantom mutation hotspots in human mitochondrial DNA

Phantom mutations are systematic artifacts generated in the course of the sequencing process. Contra common belief these artificial mutations are nearly ubiquitous in sequencing results, albeit at frequencies that may vary dramatically. The amount of artifacts depends not only on the sort of automated sequencer and sequencing chemistry employed, but also on other lab-specific factors. An experimental study executed on four samples under various combinations of sequencing conditions revealed a number of phantom mutations occurring at the same sites of mitochondrial DNA (mtDNA) repeatedly. To confirm these and identify further hotspots for artifacts, > 5000 mtDNA electropherograms were screened for artificial patterns. Further, > 30000 published hypervariable segment 1 sequences were compared at potential hotspots for phantom mutations, especially for variation at positions 16085 and 16197. Resequencing of several samples confirmed the artificial nature of these and other polymorphisms in the original publications. Single-strand sequencing, as typically executed in medical and anthropological studies, is thus highly vulnerable to this kind of artifacts. In particular, phantom mutation hotspots could easily lead to misidentification of somatic mutations and to misinterpretations in all kinds of clinical mtDNA studies.

Analysis of mitochondrial DNA variants in Japanese patients with schizophrenia

To test the hypothesis that mitochondrial DNA (mtDNA) variants contribute to the susceptibility to schizophrenia, we sequenced the entire mtDNAs from 93 Japanese schizophrenic patients. Three non-synonymous homoplasmic variants in subunit six of the ATP s

Pseudomitochondrial genome haunts disease studies

The accidental amplification of nuclear mitochondrial pseudogenes (NUMTs) can pose a serious problem for mitochondrial disease studies. This report shows that the mutation spectrum left by spurious amplification of a NUMT can be detected because it usuall

从氨基酸残基的静态可及性探讨组蛋白的分子进化

该试验利用 Apal，BamHI，BclI，BglI，BglⅡ，DraI，EcoRI，EcoRV，HaeⅡ，HindⅢ, KpnI，PstI，PvuⅡ，SacI，SalI，SmaI 和 X hoI 计18种限制性内切酶, 研究了来自欧洲、非洲及国内的5个山羊品种共计33只个体的 mtDNA, 共检测出27种限制性态型，可归结为8种单倍型. 结果表明, 国内2个山羊品种的基本单倍型为 BamHl-A, Bcl, I-B, ClaI-A, EcoRI-A, EcoRV- A, HaeIIA, HindIII-C和PvuⅡ-A, 以及为 BamHl-A,BcII-A,ClaI-A, EcoRI-A, EcoRV-A, HaeⅡ-A, HindIII-C和PvuⅡ-A.II一A, 而欧洲及非洲3个山羊品种基本的单倍型为BamHl-B, BcII-A, ClaI-A, EcoRI-A, EcoRV-A, HaeⅡ-A, HindIII-B和PvuⅡ-A，这一结果提示本研究几个受试山羊品种可能有两种不同的母系来源; 同时, 各品种的分子聚类图也表明国内2个山羊品种间具有较近的亲缘关系, 而欧洲品种和非洲3个品种间的亲缘关系较近, 这一结果也支持上述几个受试山羊品种可能有不同野生祖先的结论, 同时也验证了欧洲品种萨能羊曾引入过奴比羊的事实。

云南保种山羊线粒体DNA限制性酶切研究

　采用碱变性法,提取来自云南省不同地区4个保种山羊的13个个体的线粒体DNA(mtDNA),并用ApaⅠ,AvaⅠ,BamHⅠ,BclⅠ,BcIⅠ，BglⅡ,ClaⅠ,DraⅠ, EcoRⅠ,EcoRⅤ,HaeⅠ,HindⅢ,KpnⅠ,PstⅠ,PvuⅡ,SacⅠ,SalⅠ,SmaⅠ,StuⅠ和XhoⅠ等20种限制性内切酶进行酶切分析。结果发现它们的线粒体DNA的分子量大 小约为15.8Kb；不同限制性内切酶的酶切位点分别为：DraⅠ有7个酶切位点，AvaⅢ有6个酶切位点，EcoRⅤ和StuⅠ共有5个酶切位点，HindⅡ和HaeⅡ有4个酶 切位点，BamHⅠ,BglⅡ,PstⅠ和PvuⅡ有3个酶切位点，ApaⅠ，ClaⅠ有两个酶切位点，其余有1个酶切位点。各保种山羊间未发现变异，说明云南的4个保种山 羊极可能来自于共同的母性祖先。

一种改进的动物线粒体DNA提取方法

线粒体 DNA (mtDNA)已被广泛用于动物群体遗传学和进化生物学的研究, 并取得了许多有意义的结果. 有效的 mtDNA 提取方法无疑是开展这方面研究的前提. 关于动物 mtDNA 的提取方法, 国内外已有不少报导. 概括起来, 可分为: 1)氯化铯超速离心法, 2)柱层析法, 3)DNase法, 4)碱变性法. 该文报道了一种改进的碱变性提取法, 与其它方法相比, 具有应用范围广、简便、经济等优点。

贵州汉族、苗族、布依族和水族人群线粒体DNA多态性研究

用16种限制性内切酶对150个样本进行了mtDNA的限制性片段长度多态性(RFLP)分析。共检测到31种限制性格局, 其中Hae Ⅱ-13型、EcoRV-3型和PstⅠ-型3种限制性格局 为新报道。综合这些限制性格局, 共得出28种mtDNA类型。运用UPG法和简约法分析了各mtDNA类型之间、各人群之间的聚类关系。结果表明: 水族人群的mtDNA变异度较大; 汉族和苗族的亲缘关系最近, 布依族和水族有着较远的亲缘关系。图4表4参28

云南龙陵黄山羊线粒体DNA限制性酶切分析

采用碱性变性法提取来自于龙陵县不同地区的18只黄山羊个体的线粒体DNA(mtDNA),并用Apa I、Ava I、BamH I、Bcl I、Bgl I、Bgl II、Cla I、Dra I、EcoR I、EcoR V、Hae I、Hind III、Kpn I、Pst I、Pvu II、Sac I、Sal I、Sma I、Stu I和Xba I等20种限制性内切酶进行酶切分析。 结果发现龙陵黄山羊线粒体DNA的分子量大小约为15.8Kb;不同酶的酶切位点分别为:Dra I有7个酶切位点,Ava II有6个酶切位点,EcoR V和Stu I共有5个酶切位点,Hind III和Hea II有4个酶切位点,BamH I、Bgl II、Pst I和Pvu II有3个酶切位点,Apa I、Cla I有2个酶切位点,其余有1个酶切位点。

MITOCHONDRIAL-DNA POLYMORPHISM IN NATURAL-POPULATIONS OF DROSOPHILA-ALBOMICANS(I)

The technique of mtDNA restriction fragments length polymorphism (RFLP) was used to survey the population structure of D. albomicans. Remarkable mtDNA polymorphism has been observed in D. albomicans populations. A total of 34 nucleomorphs were detected from 82 isofemale lines assayed by only 8 restriction enzymes. The cause and the effect of this phenomenon were discussed. As a result, it is suggested that a mechanism which maintains mtDNA diversity exists in this fly, and that the high intra-populational polymorphism could numerically conceal the extent of differentiation between populations. In addition, on the base of restriction maps, it was found that the mtDNA molecule of D. albomicans might be impacted by the selection pressure during its evolution process both on the nucleotide composition and on the functional regions.

Mitochondrial genomes of two luminous beetles, Rhagophthalmus lufengensis and R-ohbai (Arthropoda, Insecta, Coleoptera)

We determined the mitochondrial MA (mtDNA) sequences of two luminous beetles (Arthropoda, Insecta, Coleoptera), Rhagophthalmus lufengensis from Yunnan, China and Rhagophthalmus ohbai from Yaeyama Island, Japan. We identified all the 37 mtDNA genes of R. l

The genetic structure and phylogenetics of pikeperch (Sander lucioperca) Anzali and Amirkolaye wetlands and perch (Perca fluviatilis) populations in Aras Dam and south-west of the Caspian Sea

The genetic structure of pikeperch (Sander lucioperca) and perch (Perca fluviatilis) populations was studied using microsatellite technique. A total of 207 specimens of adult pikeperch were collected from Aras dam (57 specimens), Anzali wetland (50 specimens), Talesh (50 specimens) and Chaboksar (50 specimens) coasts. Also a total of 158 specimens of adult perch were collected from Anzali (Abkenar (50 specimens)and Hendekhale(48 specimens)) and Amirkolaye(60 specimens) wetlands. About 2 g of each specimen's dorsal fin was removed, stored in 96% ethyl alcohol and transferred to the genetic laboratory of the International Sturgeon Research Institute. Genomic DNA was extracted using ammonium-acetate method. The quality and quantity of DNA was assessed using 1% agarose gel electrophoresis. Polymerase Chain Reaction (PCR) was conducted on the target DNA using 15 pairs of microsatellite primers. PCR products were electrophoresed on poly acryl amide gels (6%) that were stained that were stained using silver nitrate. DNA bands were analyzed with BioCapt software. Allele count and frequency, genetic diversity, expected and observed heterozygosity , allele number and the effective allele number, genetic similarity and genetic distance, Fst, Rst, Hardy Weinberg Equilibrium based on X2 and Analysis of Molecular Variance (AMOVA) at 10% confidence level was calculated using the Gene Alex software. Dendogram for genetic distances and identities were calculated using TFPGA program for any level of hierarchy. The results for P. fluviatilis showed that from 15 pair of primers that were examined 6 polymorphic and 7 monomorphic loci were produced, while 2 loci didn't produce any DNA bands. Mean allele number was 4.1±1.1 and mean observed and expected heterozygosity was 0.56±0.12 and 0.58±0.14 respectively. It was also seen that specimens from all regions were not in Hardy Weinberg Equilibrium in some of loci (P<0.001). Highest Fst (0.095) with Nm=2.37 was observed between Hendekhale and Amirkolaye and the lowest Fst (0.004) with Nm=59.31 was observed between Abkenar and Hendekhale. According to AMOVA Significant difference (P<0.05) was observed between recorded Rst in the studied regions in Anzali and Amirkolaye lagoons. In another words there are two distinct populations of this species in Anzali and Amirkolaye lagoons. The highest genetic distance (0.181) and lowest genetic resemblance (0.834) were observed between specimens from Hendekhale and Amirkolaye and the lowest genetic distance (0.099) and highest genetic 176 resemblance (0.981) were observed between specimens from Abkenar and Hendekhale. Based on the genetic dendogram tree derived by applying UPGMA algorithm, specimens from Anzali and Amirkolaye wetlands have the same ancestor. On the other hand there is no noticeable genetic distance between the specimens of these two regions. Also the results for S. lucioperca showed that from 15 pair of primers that were examined 6 polymorphic and 7 monomorphic loci were produced, while 2 loci didn't produce any DNA bands. Mean allele number was 3.0±0.6 and mean observed and expected heterozygosity was 0.52±0.21 and 0.50±0.14 respectively. It was also seen that specimens from all regions were not in Hardy Weinberg Equilibrium in some of loci (P<0.001). Highest Fst (0.093) with Nm=2.43 was observed between Aras dam and Anzali wetland and the lowest Fst (0.022) with Nm=11.27 was observed between Talesh and Chaboksar coasts. Significant differences (P<0.05) were observed between recorded Rst in the studied regions exept for Talesh and Chaboksar Coasts. In another words there are three distinct populations of this species in Caspian sea, Anzali wetland and Aras dam. Highest genetic distance (0.110) and lowest genetic resemblance (0.896) were observed between specimens from Aras dam and Anzali wetland and the lowest genetic distance (0.034) and highest genetic resemblance (0.966) were observed between specimens from Talesh and Chaboksar coasts. Based on the genetic dendogram tree derived by applying UPGMA algorithm, specimens from Talesh and Chaboksar coasts have the lowest genetic distance. On the other hand the main population of this species belongs to Anzali wetland. Phylogenetic relationship of these two species was inferred using mitochondrial cytochrome b gene sequencing. For this purpose 2 specimens of P. fluviatilis from Anzali wetland, 2 specimens of S. lucioperca from Aras dam and 2 specimens of S. lucioperca from Anzali wetland were sequenced and submitted in Gene Bank. These sequences were aligned with Clustal W. The phylogenic relationships were assessed with Mega 4. The results of evolutionary history studies of these species using Neighbor-Joining and Maximum Parsimony methods showed that the evolutionary origin of pikeperch in Aras Dam and Anzali wetland is common. On the other hand these two species had common ancestor in about 4 million years ago. Also different sequences of any region specimens are supposed as different haplotypes. 177 As a conclusion the results of this study showed that microsatellite and mtDNA sequencing methods respectively are effective in genetic structure and phylogenic studies of P. fluviatilis and S. lucioperca.

从线粒体Cyt b基因探讨红瘰疣螈物种地位的有效性

疣螈属的红瘰疣螈(Tylototriton shanjing)和棕黑疣螈(T.verrucosus)的物种界限一直不清楚.测定了来自中国西南地区14个地点的T.shanjing和T.verrucosus共40只标本的线粒体DNA Cyt b基因(753 bp).结果表明:(1)用邻接法、最大简约法和贝叶斯法等3种系统发育分析法分别重建棕黑疣螈种组系统发育树的拓扑结构不支持T.shanjing是单系群;(2)T.shanjing与T.verrucsus的mtDNA Cyt b序列差异平均值仅为1.2%.未达到种级水平.因此,全部T.shanjing样品都属于同一个物种,即T.verrucosus,不支持T.shanjing的物种地位,T.shanjing为T.verrucosus的同物异名,并建议恢复T.verrucosus的中文名红瘰疣螈.根据基于40个样品Cvt b基因序列的系统发育树和遗传变异以及地理分布,这些红瘰疣螈(T.verrucosus)样品聚为3支,即中国西南地区的红瘰疣螈可分为片马、滇中滇西和滇东南3个地理居群.

近代鸟类分类与系统发育研究进展

简要综述了宏观和微观领域的鸟类分类学研究进展。宏观领域介绍了传统形态分类学、数值分类学和支序 分类学, 结合鸣声分析强调支序分类学的应用。微观领域介绍了Sibley 分类系统和近年来出现的主要基于mtDNA 的鸟类系统学研究。通过对大量研究工作的分析提出该领域今后应进行综合性研究。

Re-examination of the phylogeny of Rhacophoridae (Anura) based on mitochondrial and nuclear DNA

The phylogenetic relationships among rhacophorid frogs are under dispute. We use partial sequences of three mitochondrial (12S rRNA, 16S rRNA, and cytochrome b) and three nuclear protein-coding (Rag-1 rhodopsin exon 1, and tyrosinase exon 1) genes from 57

Molecular and morphological analysis of endangered species Gymnodiptychus integrigymnatus (Teleostei: Cyprinidae)

Gymnodiptychus integrigymnatus is a critically endangered species endemic to the Gaoligongshan Mountains. It was thought to be only distributed in several headwater-streams of the Longchuanjiang River (west slope of the Gaoligongshan Mountains, belonging to the Irrawaddy River drainage). In recent years, dozens of G. integrigymnatus specimens have been collected in some streams on the east slope of the Gaoligongshan Mountains (the Salween drainage). We performed a morphological and genetic analyses (based on cytochrome b and D-loop) of the newly discovered populations of G. integrigymnatus to determine whether the degree of separation of these populations warrants species status. Our analysis from the cytochrome b gene revealed that nine individuals from the Irrawaddy drainage area and seven individuals from the Salween drainage area each have only one unique haplotype. The genetic distance between the two haplotypes is 1.97%. Our phylogenetic analysis revealed that G. integrigymnatus is closely related to highly specialized schizothoracine fishes. Analysis from the mitochondrial control region revealed that G. integrigymnatus has relatively high genetic diversity (pi was 0.00891 and h was 0.8714), and individuals from different river drainages do not share the same haplotypes. The AMOVA results indicated 87.27% genetic variability between the Salween and Irrawaddy populations. Phylogenetic trees show two major geographic groups corresponding to the river systems. We recommend that G. integrigymnatus should be considered as a high priority for protected species status in the Gaoligongshan Mountains National Nature Reserve, and that the area of the Gaoligongshan Mountains National Nature Reserve should be expanded to cover the entire distribution of G. integrigymnatus. Populations of G. integrigymnatus from different river systems should be treated as evolutionarily significant units.