中国稻瘟病菌的致病性和其DNA指纹图谱间关系的研究

通过在稻瘟病菌Pyricularia oryza。基因组文库中的筛选,找到一个散布的并具有基因组特异性的层重复顺序POR6。本文报道用这一株针对6个日本菌株和26个中国北方菌株进行DNA指纹作图的结果。其中22个中国北方菌株按其杂交带型百分相似率被分成8个株系。一些在我们实验室保存的菌株用传统方法鉴定发现在转管过程中会发生致病性变异。当用POR6作探针与这些菌株NNA的EcoRV酶切片段杂交时,检测出它们的无性世代中出现数条EcoRV多态性片段。     

真核细胞重复顺序DNA研究的一些进展

本文从如下几方面介绍了真核细胞重复顺序DNA结构功能研究的一些进展:(1)串联重复顺序是构成基因多态性的分子基础,基因多态性分析已用于家系分析;(2)一些短的重复顺序如LTR上具有转录调节信号,对基因表达起调控作用;(3)一些重复顺序的表达,与细胞生长分化有一定的相关性;(4)重复顺序在种属演化中具有一定的意义。     

稻瘟病菌株的DNA指纹分析及小种鉴别

基于对稻瘟病菌(Pyricularia oryzae)基因文库的分析,我们找到了一套含重复顺序的克隆。其中POR6和POR7被证实具有高度的多态性并随机散布于稻瘟病菌生理小种的致病性时,可以获得可分辨的基因组特异的杂交带型。我们还分析了致病性与8个稻瘟病菌株DNA指纹图谱之间的关系,结果表明各个小种组合间的百分相似率S_xy,值与该小种组合间共同侵染的鉴别品种数目有正相关性。     

刺猬高重复顺序DNA的分子克隆及核苷酸顺序分析

为了分析比较食虫目高重复顺序的结构特征,我们将食虫目动物刺猬的高重复顺序DNA进行了分离、纯化、分子克隆,并测出了1186bp的核苷酸顺序。经分析发现在碱基组成上它与食虫目另一动物臭鼩有许多相似之处,如:G-C碱基对与A-T碱基对在整个片段中分布很不均衡;片段内部含有许多连续多次重复的短片段等,很可能为食虫目动物所特有。     

一些水稻的重复DNA顺序在稻属和其它禾本科植物中的多态性

重复DNA顺序是真核生物基因组的特征,很多重复DNA顺序已从小麦、拟南芥菜、燕麦、水稻、玉米等植物的基因组中克隆出来,还发现有一些重复DNA顺序具有基因组特异性,用它们作探针可以分析同属或同科物种的起源和亲缘关系,并建立系统进化树。小卫星DNA或微小卫星DNA所产生的指纹图谱可作为一种遗传学标志来研究系统进化、染色体的精细结构和物种的鉴定。一些中度重复DNA序列还可以作为组织培养株系和细胞杂交筛选的分子标志。稻属已发现并定名的有22个种,根据杂交亲和性、细胞遗传学和生理生化等将它分为6个二倍体组型(AA、BB、CC、DD、EE和FF)和2个四倍体组型(BBCC和CCDD)。现多把禾本科分作5个亚科:竹亚科、稻亚科、早熟禾亚科、画眉草亚科和     

水稻(Oryza sativa L.)重复DNA顺序的克隆及序列分析——重复顺序存在意义的探讨

以大肠杆菌启动子选择质粒pKK175-6、pKK232-8为载体,将通过复性动力学方法获得的水稻重复DNA顺序片段进行克隆。阳性克隆菌株表现出对四环素或氯霉素不同程度的抗性。DNA序列分析表明,克隆到的某些水稻重复DNA顺序具有丰富的基因表达调节元件的同源序列。     

重复DNA顺序pRRD9在稻属中的拷贝数测定

本文应用狭缝印渍杂交方法,把水稻基因组总DNA和含水稻中度重复顺序片段的质粒(pRRD9)DNA分别转移到尼龙膜上形成狭缝印渍、然后用~(32)P标记的 pRRD9插入片段进行杂交、根据各狭缝印渍的放射性强度,测定水稻(Oryza)一些栽培种和野生种基因组中重复DNA顺序的拷贝数,并就拷贝数与水稻进化关系及基因组型的联系进行讨论.     

水稻重复DNA顺序克隆pRRD3的研究

用ＤＮＡ复性动力学方法克隆到一个水稻（ＯｒｙｚａｓａｔｉｖａＬ．）中度重复ＤＮＡ顺序。不同限制性内切酶消化和Ｓｏｕｔｈｅｒｎ杂交分析显示,这段重复ＤＮＡ顺序以串联加散布的形式存在于水稻基因组中;序列分析表明在它内部含有一个典型的植物启动子序列ＴＧＴＡＴＡＡＡＴＡ;以ｐＲＲＤ３克隆片段作探针,对水稻３４个品种进行拷贝数测定,在野生稻与栽培稻、籼稻与粳稻之间均存在拷贝数上的明显差异;对ＡＡ基因组不同亚型水稻ＤＮＡ进行Ｓｏｕｔｈｅｒｎ杂交分析,得到基因组亚型特异的杂交带谱,说明该重复顺序是研究水稻进化和分类的一个有用探针。     

水稻染色体DNA的交变脉冲电泳分析

从水稻完整的细胞核中释放出来的染色体DNA在交变脉冲电泳申表现为一种“240kb单位”的形式。这种单位,经限制性内切酶消化,可产生连续分布的大至1500kb的染色体DNA片段。文章讨论了“240kb单位”作为水稻染色体DNA基本结构单位的可能性。     

Assignment of Alu-repetitive sequences to large restriction fragments from human chromosomes 6 and 22

We have employed a pulsed field gel electrophoresis and Alu hybridization approach for identification of large restriction fragments on chromosome 6 and 22. This technique allows large portions of selected human chromosomes to be visualized as discrete hybridization signals. Somatic cell hybrid DNA which contains chromosome 6 or chromosome 22 was restricted with either Notl or Mlul. The restriction fragments were separated by pulsed field gel electrophoresis (PFGE) and hybridized against an Alu repetitive sequence (Blur 8). The hybridization signals result in a fingerprint-like pattern which is unique for each chromosome and each restriction enzyme. In addition, a continuous pattern of restriction fragments was demonstrated by gradually increasing puls times. This approach will also be suitable to analyze aberrant human chromosomes retained in somatic cell hybrids and can be used to analyze flow sorted human chromosomes. To this end, our method provides a valuable alternative to standard cytogenetic analysis.     

交变脉冲电场凝胶电泳与植物大分子DNA的制备

介绍了交变脉冲电场凝胶电泳的原理、方法及其在植物大分子ＤＮＡ制备方面的应用     

DNA fingerprinting of human isolates of Salmonella enterica serotype Paratyphi B in Malaysia

AIMS: DNA fingerprinting of Salmonella enterica serotype Paratyphi B isolated in Malaysia during 1982-83, 1992 and 1996-2002 was carried out by pulsed-field gel electrophoresis (PFGE), antimicrobial susceptibility tests and D-tartrate utilization tests to assess the extent of genetic diversity of these isolates in Malaysia. METHODS AND RESULTS: Eighty-six human isolates and one food isolate of Salm. Paratyphi B were analysed by PFGE, antimicrobial susceptibility tests and D-tartrate utilization tests. Sixty-five strains were D-tartrate-negative (dT-) while 22 strains were D-tartrate-positive (dT+). Thirty-seven per cent of the Salm. Paratyphi B strains were resistant to one or more antimicrobial agents. PFGE analysis clearly distinguished the dT- and dT+ strains into two clusters based on the unweighted pair group average method (UPGMA). Twenty-two XbaI-pulsotypes were observed among the 65 dT- strains while 17 XbaI-pulsotypes were observed among the 22 isolates of Salm. Paratyphi B dT+. CONCLUSIONS: The present study showed that PFGE was very discriminative with 33.7% of the strains yielding distinct fingerprints. Paratyphoid fever in Malaysia is probably caused by one predominant, endemic clone of Salm. Paratyphi B dT- with various subtypes. There was no association between the pulsotypes and the severity of the disease indicating that the severity of the disease is probably multifactorial. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings of the present study verify the usefulness of PFGE in characterizing strains of Salm. Paratyphi B. This is the first report on the application of PFGE on a large collection of Salm. Paratyphi B in Malaysia.     

检测植物DNA扩增多态性方法的比较和改进

以辽东栎(Quercus liaotungensis Koidz.)、锦鸡儿(Cargagana ssp.)和野大豆(Glycine soja(L.)Sieb.etZucc.)为材料,比较了随机扩增多态DNA(RAPD)和DNA扩增指纹(DAF)方法。用RAPD的琼脂糖胶电泳和溴乙锭染色,RAPD和DAF谱一般不足10条带。用DAF的变性聚丙烯酰胺凝胶电泳(PAGE)和银染,极大地提高了RAPD的灵敏度和分辨率,多达20～40个产物。用3＇末端完全相同的引物,RAPD和DAF有同样的扩增谱,说明两种方法有相似的机理。降低胶的浓度可提高RAPD和DAF的分辨率,达40～80条带。琼脂糖电泳分离的溴乙锭显示的单荧光带,用PAGE和银染可分辨出多个片段。分子克隆证实单荧光带的分子量异质性。在用Taq DNA多聚酶的条件下,RAPD和DAF的再现性均良好。     

凝胶条带内DNA回收测序评估DGGE可靠性

为了评估DGGE的可靠性,对DGGE条带中回收的DNA片段进行了测序比较分析,并引入了DGGE可靠性指数的概念评价其可靠性。结果显示同一条DGGE条带回收的DNA来自同一属的概率为64.7%,相同位置的DGGE条带可以被认为是同一OTU;不同的DGGE条带回收到类似的DNA序列（16S rDNA V3区差异小于4 bp）的概率为10.5%;DGGE可靠性指数为74.8%。以上结果表明尽管DGGE技术与理论预期存在一定的差距,但是DGGE技术基本能够反映微生物群落的多样性。       

The use of (GTG)5 oligonucleotide as an RAPD primer to type Campylobacter concisus

AIM: DNA fingerprinting using (GTG)(5) oligonucleotide as a primer in a random amplified polymorphic DNA (RAPD) assay was assessed by typing isolates of Campylobacter concisus strains, collected over a period of 8 years. METHODS AND RESULTS: RAPD analysis using the (GTG)(5) oligonucleotide as a primer was used to type 100 isolates of C. concisus comprising mostly isolates from children with diarrhoea. Using this method, 86% of the isolates were found to be genotypically diverse. Of these heterogeneous isolates, 25 of the strains were also shown to be genetically distinct in a previous study using pulsed field gel electrophoresis. The remaining isolates (14) could be classified into five profile groups based on the DNA fingerprinting patterns. The assay successfully identified epidemiologically linked strains from the unrelated genetically diverse pool of strains. CONCLUSIONS: Laboratory RADP typing using the (GTG)(5) primer proved to be useful in distinguishing related strains of C. concisus from a large pool of unrelated strains of this organism. SIGNIFICANCE AND IMPACT OF THE STUDY: RAPD typing using (GTG)(5) is a simple method that could be used to investigate the epidemiology of C. concisus. The results suggest that homologous lineages of C. concisus may exist within an otherwise heterogeneous species complex. However, these data need to be confirmed using a more robust typing method.     

金鱼(红龙睛)次黄嘌呤-鸟嘌呤磷酸核糖转移酶的分离及特性初步研究

 从金鱼肝脏分离并部分纯化了HGPRT。初步结果表明,该酶的分子量为78,000左右。由3个亚基组成,每个亚基的分子量约为27,000。金鱼的HGPRT很稳定,在85℃加热10分钟,至少保留80％的酶活性。该酶与PRPP的反应产物在淀粉凝胶电泳图谱上出现两条反应带。     

不同DGGE谱带信息提取方法对分析结果的影响

自1993年Muyzer,et al.[1]将变性梯度凝胶电泳技术(Denaturing gradient gel electrophoresis,DGGE)引入到微生物生态学研究以来,DGGE已被广泛应用于各种生态系统(如淡水、海洋、土壤、动物消化道等)的微生物群落结构分析[2—6]。但对于DGGE凝胶的分析至今仍没有统一的方法,