微卫星DNA技术及其应用的探讨

微卫星DNA是一类短串联重复的寡核苷酸，广泛的分散于整个基因组中，具有多态性高、突变速率快等特点，在医学领域、生物学领域应用广泛。本文综述了微卫星多态性形成机制、序列获得方法及其应用途径，尤其是微卫星不稳定性与疾病的相互关系。     

SSR分子标记在作物遗传多样性研究中的应用现状

微卫星(microsatellites)或简单序列重复(simple sequence repeats,SSRs)是以1-6个碱基为基本单元的串联重复序列,具有含量丰富、多态性高、共显性和检测方法简单等优点,是研究植物遗传多样性的主要的分子标记方法之一,已被用于多种农作物的遗传多样性研究。概述了微卫星DNA标记的原理及特点,探讨了其在农作物物遗传多样性研究中的应用和发展前景。     

DNA分子标记与动物育种

论述了DNA分子标记的分类以及限制性酶切片段长度多态性、随机扩增多态性DNA、扩增片段长度多态性、简单重复序列、单核苷酸多态性等5种主要DNA分子标记的基本原理和优缺点。同时，还分析了它们在动物遗传育种中应用情况和发展前景。     

金发藓目系统学研究概况

结合经典分类学和同工酶、等位酶分析技术、随机扩增多态性DNA、限制性片段长度多态性、简单重复序列和DNA序列测序等分子生物学技术对金发藓目植物的系统学研究概况进行了介绍．并指出了在金发藓目植物系统学研究中存在的一些问题。     

DNA指纹图技术及其在实验动物学中的应用

实验动物要求具有明确的遗传背景 ,实验动物的遗传质量直接影响生物医学研究结果的准确性、重复性与科学性 ,遗传质量监测是保证实验动物遗传质量的重要措施[1] 。现在实验动物的遗传质量监测运用生化标记等表型检测方法 ,存在灵敏度低等局限性。随着分子生物学技术的发展 ,DNA分子标记技术的出现 ,为实验动物遗传物质DNA的直接检测带来了新的可行的技术方法。1　DNA指纹技术的原理真核生物的基因组大且复杂 ,广泛分布着各种形式的重复序列 ,由于它们不编码肽链 ,很少受到自然选择作用的影响而保持着高度的多态性。这些重复序列按其在染…     

DNA序列高维空间数字编码运算法则的进一步结果

研究了DNA序列高维空间数字编码的更一般的运算法则:充分利用陈惟昌等人提出的DNA序列高维空间的表观维数Nv,数值维数Nx以及差异维数Nd,讨论了当Nd=0,1,2,2n或2n+1(n=0,1,2,…)时,具体刻画了DNA序列的首段碱基及其数值取值范围;推导出DNA序列多点突变(单核苷酸多态性SNP)的运算法则;利用DNA序列的定值部Xi和定位部Qi及其计算公式,从新的角度导出DNA重复序列的编码法则和运算法则.     

着丝粒串联重复序列RCS2对亚洲栽培稻和非洲栽培稻的比较分析

为了揭示中高度重复序列在同为AA基因组的亚洲栽培稻和非洲栽培稻基因组中的差异以及重复序列在.栽培稻种的分化过程中可能起到的作用,利用水稻着丝粒串联重复序列RCS2作为探针分别对籼稻广陆矮4号、粳稻日本晴和非洲栽培稻的体细胞染色体进行荧光原位杂交(FISH)实验,并对其核型进行同源性聚类和比较分析,杂交结果显示:RCS2序列位于在3种栽培稻染色体组中,RCS2序列位于每条染色体的着丝粒位置,但有不同的分布特点,表明该3种栽培稻基因组的RCS2序列有不同的进化方向.探讨了RCS2序列结合Cot-1 DNA FISH方法对水稻染色体组进行核型分析的可行性和优势.     

微卫星标记在Ⅱ型糖尿病基因定位上的应用

微卫星是以几个碱基(一般为1～6个)为重复单位组成的简单的串联重复序列,遍布真核生物基因组中,具有丰度高、多态性高、共显性标记、选择中性、可自动检测等优点,已经成为人类遗传学研究中被广泛应用的分子遗传标记.Ⅱ型糖尿病是糖尿病中最普遍的一种形式,它以胰岛素抗性活动紊乱为特征.该病与遗传因素、肥胖以及缺少运动行为等生活方式相关.为有效阻止或至少延缓疾病的进程,对Ⅱ型糖尿病的基因定位研究是必要的.微卫星DNA标记技术在动物亲缘关系的鉴定、特定基因定位、群体遗传结构的分析、物种的进化和系统发生以及遗传基因连锁图谱的构建等方面已得到了广泛的应用.因此,就微卫星DNA遗传标记在Ⅱ型糖尿病基因定位上应用的原理、方法等方面进行综述.     

浅蛤属(Macridiscus)物种特异性线粒体DNA分子标记及其长度多态性和个体内异质性

浅蛤属(Macridiscus)包括3个形态非常相似的物种,即黑浅蛤(M .melanaegis)、等边浅蛤(M .multifarius)和半步目浅蛤(M .semicancellata),其中前二者为同域分布.根据形态学特征,浅蛤属物种较难区分.本研究以改进的CTAB法提取浅蛤个体基因组DNA 后,用引物Macr_CO1_F16834和Cytb_Macr_R2PCR 配对进行聚合酶链式反应(PCR),扩增线粒体DNA控制区至细胞色素b基因(Cytb)的DNA 片段并进行序列测定,结果发现在黑浅蛤控制区偏3′端,存在该种特有的长度为363bp的数目可变串联重复序列(variablenumberoftandemrepeat,VNTR),种内因该VNTR出现1~3次不等的串联拷贝,而表现出广泛的长度多态性和个体内线粒体异质性现象,这是目前在动物线粒体控制区中发现的最大的VNTR;在半步目浅蛤的控制区的偏3′端,则同时存在14和17bp的2个较短的VNTRs,该物种也因这2个VNTRs的拷贝数目不同,而存在广泛的长度多态性;而等边浅蛤的该段序列长度保守,没有与其他物种共享的VNTR.以该段序列作为分子标记,运用PCR技术能够快速准确地识别浅蛤属物种.      

微卫星DNA在亲权关系鉴定中的应用

微卫星DNA是广泛分布的,具有高度多态性的分子标记.适用于进行亲权关系鉴定、系统发育、遗传多样性等研究.主要介绍了利用微卫星DNA进行亲权关系鉴定的原理与方法以及研究最新进展.     

Individual-specific 'fingerprints' of human DNA

Simple tandem-repetitive regions of DNA (or 'minisatellites') which are dispersed in the human genome frequently show substantial length polymorphism arising from unequal exchanges which alter the number of short tandem repeats in a minisatellite. We have shown previously that the repeat elements in a subset of human minisatellites share a common 10-15-base-pair (bp) 'core' sequence which might act as a recombination signal in the generation of these hypervariable regions. A hybridization probe consisting of the core repeated in tandem can detect many highly polymorphic minisatellites simultaneously to provide a set of genetic markers of general use in human linkage analysis. We now show that other variant (core)n probes can detect additional sets of hypervariable minisatellites to produce somatically stable DNA 'fingerprints' which are completely specific to an individual (or to his or her identical twin) and can be applied directly to problems of human identification, including parenthood testing.     

Minisatellite repeat coding as a digital approach to DNA typing

Most DNA typing systems used in forensic and legal medicine assay allelic length variation at tandem repetitive DNA regions such as minisatellites. A simple alternative approach that displays patterns of variant repeat units along minisatellite alleles is described here. This produces DNA profiles as extraordinarily variable digital sequences appropriate for forensic investigations, including computer databasing, and for analysing allele diversity and the role of recombination in minisatellite instability.     

Isolation and identification of a novel STR (D17S2204) in the vicinity of NF1 locus

Using a synthetic oligonucleotide (dA-dC)15 as a probe to screen a probe pool made by microdissected human chromosome band 17q11-q12, a DNA fragment containing (CA)16 was isolated, which is a novel short tandem repeat (STR) determined by searching in the GenBank and GDB. It has 8 allelic types with a PIC value of 0.61. The novel STR is conformed with Mendelian segregation according to the linkage analysis of a three-generation neurofibromatosis type Ⅰ (NF1) pedigree. The STR was localized at chromosome band 17q11-q12 in the vicinity of NF1 gene by FISH analysis. The accession number of the STR in GenBank and GDB is G32112 and D17S2204 respectively.     

辽南地区汉族人群Penta E和Penta D基因座的多态性分析

对辽南地区汉族群体的两个短串联重复(Short tandem repeats STR)基因座等位基因频率进行研究，得到辽南地区汉族群体Penta E和Penta D基因座的群体遗传学数据．EDTA抗凝血采自172名无血缘关系的汉族个体，采用Celex-100法抽提DNA，荧光标记PCR扩增，310型全自动测序仪电泳检测．得到了Penta E和Penta D基因座的等位基因频率，Penta E和Penta D基因座等位基因的杂合度分别为0．8846和0．8078；个人识别率分别为0．9782和0．9326．两个STR基因座具有较高的杂合度和个人识别能力，等位基因分布符合Hardy-Weinberg平衡，是理想的遗传标记，可用于法医学个体识别和亲权鉴定。     

DNA sequences of telomeres maintained in yeast

Telomeres, the ends of eukaryotic chromosomes, have long been recognized as specialized structures. Their stability compared with broken ends of chromosomes suggested that they have properties which protect them from fusion, degradation or recombination. Furthermore, a linear DNA molecule such as that of a eukaryotic chromosome must have a structure at its ends which allows its complete replication, as no known DNA polymerase can initiate synthesis without a primer. At the ends of the relatively short, multi-copy linear DNA molecules found naturally in the nuclei of several lower eukaryotes, there are simple tandemly repeated sequences with, in the cases analysed, a specific array of single-strand breaks, on both DNA strands, in the distal portion of the block of repeats. In general, however, direct analysis of chromosomal termini presents problems because of their very low abundance in nuclei. To circumvent this problem, we have previously cloned a chromosomal telomere of the yeast Saccharomyces cerevisiae on a linear DNA vector molecule. Here we show that yeast chromosomal telomeres terminate in a DNA sequence consisting of tandem irregular repeats of the general form C1-3A. The same repeat units are added to the ends of Tetrahymena telomeres, in an apparently non-template-directed manner, during their replication on linear plasmids in yeast. Such DNA addition may have a fundamental role in telomere replication.     

串联表达载体中短重复序列(BmKb1)_n模板的PCR效率及其优化条件研究

采用Overlap PCR法获取BmKb1基因,构建载体pGEX-6p-1/BmKb1,同尾酶技术用于构建串联表达载体pGEX-6p-1/(BmKb1)n(n=2,4,6,8,10)。以pGEX-6p-1/(BmKb1)n为模板,使用标准PCR程序研究短重复序列PCR效率;以BmKb1八倍体为模板,分别研究PCR循环数、引物终浓度及退火温度对短重复序列PCR效率及特异性的影响。结果表明:串联重复序列数目增加,则非特异性条带增加,特异性条带含量减少;BmKb1基因串联体PCR扩增在18～20循环数、引物终浓度为0.05～0.1μM、60～62℃较高退火温度时,可获得相对高产量高特异性BmKb1基因串联体PCR片段。本研究将为串联表达载体克隆、筛选、测序及基因组STRs序列克隆测序提供技术参考。     

十拷贝串联重复laGLP-1基因植物表达载体的构建及对黄瓜遗传转化的研究

天然胰高血糖素样肽-1(glucagon-like peptide-1,GLP-1)是含有30个氨基酸的短肽激素,对2型糖尿病具有特殊疗效.以本实验室克隆的长效GLP-1(long acting GLP-1,laGLP-1)基因为基础,构建了十拷贝串联重复laGLP-1的无抗生素选择标记的植物表达载体pX6-laGLP-1,采用农杆菌介导法对黄瓜自交系2M1进行转化,通过PCR及Southern杂交检测分析,最后获得了4株稳定整合十拷贝串联重复laGLP-1的转基因植株.该遗传体系的建立为研发糖尿病食疗型转基因植物或用其生产治疗糖尿病的口服药物奠定了基础.     

Analysis of tandem repeats in the genome of Chinese shrimp Fenneropenaeus chinensis

Through random sequencing, we found a total of 884000 base-pairs （bp） of random genomic sequences in the genome of Chinese shrimp （Fenneropenaeus chinensis）.Using bio-soft Tandem Repeat Finder （TRF） software, 2159 tandem repeats were found, in which there were 1714 microsatellites and 445 minisatellites, accounting for 79.4% and 20.6% of repeat sequences, respectively. The cumulative length of repeat sequences was found to be 116685 bp, accounting for 13.2% of the total DNA sequence; the cumulative length of microsatellites occupied 9.78% of the total DNA sequence, and that of minisatellites occupied 3.42%. In decreasing order, the 20 most abundant repeat sequence classes were as follows： AT （557）, AC （471）, AG （274）, AAT（92）, A （56）, AAG （28）, ATC （27）, ATAG （27）, AGG （18）, ACT（15）, C （11）, AAC （11）, ACAT （11）, CAGA （10）, AGAA （9）,AGGG （7）, CAAA （7）, CGCA （6）, ATAA （6）, AGAGAA （6）.Dinucleotide repeats, not only in the aspect of the number,but also in cumulative length, were the preponderant repeat type. There were few classes and low copy numbers of repeat units of the pentanucleotide repeat type, which included only three classes： AGAGA, GAGGC and AAAGA. The classes and copy numbers of heptanucleotide, eleven-nucleotide and thirteen-nucleotide primer-number-composed repeats were distinctly less than that of repeat types beside them.