多重荧光PCR同时检测转基因成分35S和Nos方法的建立

根据商品化转基因作物中常用的花郴花花叶病毒启动子（CaMV35S）和根癌农杆菌终止（Nos）的序列特点，设计并合成了两对引物和相对应的荧光双链探针，建立一种应用荧光双链探针的多重荧光PCR同时检测转基因成分35S启动子和Nos终止子的方法，并利用该方法对马铃薯、大豆、玉米、甜椒、番茄等11份实物样品进行了检测，其中有5份样品结果阳性，结果表明所建立的多重荧光PCR方法能同时检测了35S方法同时检测出35S和Nos双组分，较常规PCR技术更为简便、快速、准确，有很很好的应用前景。     

A DNA sequence alignment algorithm using quality information and a fuzzy inference method

DNA sequence alignment algorithms in computational molecular biology have been improved by diverse methods. In this paper, we propose a DNA sequence alignment that uses quality information and a fuzzy inference method developed based on characteristics of DNA fragments and a fuzzy logic system in order to improve conventional DNA sequence alignment methods that uses DNA sequence quality information. In conventional algorithms, DNA sequence alignment scores are calculated by the global sequence alignment algorithm proposed by Needleman-Wunsch, which is established by using quality information of each DNA fragment. However, there may be errors in the process of calculating DNA sequence alignment scores when the quality of DNA fragment tips is low, because only overall DNA sequence quality information are used. In our proposed method, an exact DNA sequence alignment can be achieved in spite of low quality of DNA fragment tips by improvement of conventional algorithms using quality information. Mapping score parameters used to calculate DNA sequence alignment scores are dynamically adjusted by the fuzzy logic system utilizing lengths of DNA fragments and frequencies of low quality DNA bases in the fragments. From the experiments by applying real genome data of National Center for Biotechnology Information, we could see that the proposed method is more efficient than conventional algorithms.     

基于聚类分析的DNA序列分类研究

从DNA序列片段的个案中密码子分布密度角度出发,提取出DNA序列片段的特征,应用模糊数学中的模糊聚类分析理论对DNA序列片段进行分类．由DNA序列片段中64种密码子出现的频率,给出两个案夹角余弦的定义,由两个案的交角余弦来描述个案之间的相关性．采取分层聚类分解法,应用SPSS统计软件,计算出描述个案之间相关性的模糊矩阵,同时给出DNA序列片段的分类结果．仿真结果表明,该算法具有分类简单且分类结果精度高的优点．     

A DNA sequence alignment algorithm using quality information and a fuzzy inference method

DNA sequence alignment algorithms in computational molecular biology have been improved by diverse methods. In this paper, we propose a DNA sequence alignment that uses quality information and a fuzzy inference method developed based on the characteristics of DNA fragments and a fuzzy logic system in order to improve conventional DNA sequence alignment methods that uses DNA sequence quality information. In conventional algorithms, DNA sequence alignment scores are calculated by the global sequence alignment algo- rithm proposed by Needleman-Wunsch, which is established by using quality information of each DNA fragment. However, there may be errors in the process of calculating DNA sequence alignment scores when the quality of DNA fragment tips is low, because only the overall DNA sequence quality information are used. In our proposed method, an exact DNA sequence alignment can be achieved in spite of the low quality of DNA fragment tips by improvement of conventional algorithms using quality information. Mapping score param- eters used to calculate DNA sequence alignment scores are dynamically adjusted by the fuzzy logic system utilizing lengths of DNA fragments and frequencies of low quality DNA bases in the fragments. From the experiments by applying real genome data of National Center for Biotechnology Information, we could see that the proposed method is more efficient than conventional algorithms.     

融合智能检测的DNA序列预处理新方法

提出一种融合智能检测的DNA序列预处理新方法。该方法不需要预先给出载体序列、剪接位点和克隆适配片段等信息,通过统计分析、随机搜索和构建图操作等方法自动发现并定位垃圾信息。以Zebrafish DNA序列为样本进行的预处理实验结果证明该方法能够显著提高DNA序列预处理的效率和准确性,在处理超长序列时更稳定、错误率更低。     

用于DNA序列结构分析的特征抽取方法

人类基因组计划中的DNA序列图谱是生命科学和基因工程中的伟大成就,要解译隐藏在基因组中的生物信息还有一段很长的路要走,这是因为DNA序列的结构很难分析和识别．作者提出一种用于DNA序列结构分析的特征抽取方法．这种方法采用DNA序列码序的共生概率来抽取高维特征．然后采用相关法和／或贝叶斯分类器来分类结构模式．一些仿真试验的结果表明这种方法适合于DNA序列的结构分析。     

Kas基因启动子区的克隆及功能初步分析

利用TAIL-PCR技术,克隆到了与辣椒素合成有关的胎座特异表达基因——3-酮酯酰．ACP合成酶基因（Kas）上游400bp的调控区域．将其全长片段与GUS基因连接构建植物表达载体并转化烟草．GUS组织化学染色表明,克隆到的440bp片段具有启动子活性．对该片段进行序列分析发现,在起始密码子ATG上游存在2个TATA-box,分别为-316～-311位的TATAAA和-224～-219位的TATAAA;在TATA-box上游还存在1个位于-378～-374处的CAAT-box,序列为CCAAT．该研究旨在为利用基因调控辣椒素的生物合成,提高辣椒果实中的辣椒素含量奠定基础．     

过氧化草酸酯化学发光检测DNA荧光探针

根据过氧化草酸酯与双氧水作用产生的高能中间体激发荧光物质发光的原理,研究了利用过氧化草酸酯TCPO-H2O2化学发光体系测定Cy5标记的DNA探针的新方法,探讨了溶剂、酸度、催化剂及发光溶液浓度对化学发光信号的影响,确立了最佳分析条件.其线性范围为8.4×10-9~2.8×10-6mol/L,检出限(3σ)为2.8×10-9mol/L.据此建立了操作简便、灵敏度高的测定Cy5标记的DNA探针的新方法.     

富钾植物DNA导入早稻变异后代的RAPD分析

应用花粉管通道法和浸种法将两种富钾植物空心莲子草和商陆DNA导入到不同的水稻品种中,结果在其后代产生了广泛的变异,并从中筛选出耐低钾的变异材料。应用PCR技术对其中耐低钾的4个变异材料进行了RAPD分析。结果表明,所用100个随机引物中有7个引物在受体和变异后代间扩增出了多态性产物。后代和受体的相似系数均在90%以上,而与供体的相似系数只有6%以下,其中有三个引物的扩增产物中出现了供体的特异性带。这说明空心莲子草和商陆DNA导入水稻后确实引起了水稻基因组结构的变化,同时也为变异后代所发生的生物学性状的改变提供了直接的分子证据。     

Discovery and demonstration of small circular DNA molecules derived from Chinese tomato yellow leaf curl virus

Tomato yetlow leaf curl viruses betong to Begomoviruses of geminiviruses. In this work, we first found and demonstrated that the small circular DNA molecules were derived from Chinese tomato yetlow leaf curl viruses (TYLCV-CHI). These small circular DNA molecules are about 1,3 kb, which are half the full-length of TYLCV-CHI DNA A. It was shown by sequence determination and analysis that there was unknown-origin sequence insertion in the middle of the small molecules. These sequences of unknown-origin were neither homologous to DNA A nor to DNA B, and were formed by recombination of virus DNA and plant DNA. Although various defective molecules contained different unknown-origin sequence insertion, all the molecules contained the intergenic region and part of the AC1 (Rep) gene. But they did not contain full ORF.     

DNA序列分析在药用植物鉴定中的应用

随着DNA测序技术的不断发展,DNA序列分析在药用植物鉴定和品质研究等方面得到了广泛应用,为药用植物鉴定和道地性研究注入了新的活力.从药用植物鉴定和品质研究中的常用DNA片段(基因)、DNA序列分析在药用植物鉴定以及道地性研究中的应用等3个方面对有关研究的主要进展进行了综述,并对今后研究的前景和应注意的问题进行了初步讨论.     

Positive identification of an immigration test-case using human DNA fingerprints

The human genome contains a set of minisatellites, each of which consists of tandem repeats of a DNA segment containing the 'core' sequence, a putative recombination signal in human DNA. Multiallelic variation in the number of tandem repeats occurs at many of these minisatellite loci. Hybridization probes consisting of tandem repeats of the core sequence detect many hypervariable minisatellites simultaneously in human DNA, to produce a DNA fingerprint that is completely individual-specific and shows somatic and germline stability. These DNA fingerprints are derived from a large number of highly informative dispersed autosomal loci and are suitable for linkage analysis in man, and for individual identification in, for example, forensic science and paternity testing. They can also be used to resolve immigration disputes arising from lack of proof of family relationships. To illustrate the potential for positive or inclusive identification, we now describe the DNA fingerprint analysis of an immigration case, the resolution of which would have been very difficult and laborious using currently available single-locus genetic markers.     

含取代基的金属酞菁的合成与光伏效应

合成了含取代基的酞菁铜，如十六氯酞菁铜、十六氢酞菁铜、四硝基酞菁铜及四氨基酞菁铜等．测定了这些衍生物在铂电极上和砷化镓电极上的光伏效应．结果表明含拉电子取代基的酞菁铜比含推电子取代基的酞菁铜的光伏效应大．     

地中海伞藻叶绿体生物节律基因的分子克隆及限制酶图谱分析

本用分离纯化的地中海伞藻叶绿体ＤＮＡ，以ｐｃｏｓ２ ＥＭＢＬ为载体构建了ｃｏｓｍｉｄ分子克隆库。并用果蝇的与生物节律控制相关的ＤＮＡ顺序为探针，从地中海伞藻叶绿体ｃｏｓｍｉｄ分子克隆库中筛选到一组含有同源顺序的克隆，经印迹杂交和限制酶分析，这些克隆都含有与果蝇生物节律探针强烈同ＤＮＡ顺序，且各克隆的限制酶图谱都相互重叠。这一结果表明，在地中海伞藻叶绿体基因组中可能也存在与果蝇相同或基本相同的与生     

Cloning and sequence analysis of a gene encoding polygalacturonase-inhibiting protein from cotton

Polygalacturonase-inhibiting proteins (PGIP) play important roles in plant defense of pathogen, especially fungi. A pair of degenerated primers is designed based on the conserved sequence of 20 other known pgip genes and used to amplify Gossypium barbadense cultivation 7124 cDNA library by touch-down PCR. A 561 bp internal fragment of the pgip gene is obtained and used to design the primers for rapid amplification of cDNA ends. A composite pgip gene sequence is constructed from the products of 5′ and 3′ RACE, which are 666 bp and 906 bp respectively. Analysis of nucleic acid sequence shows 69.2% and 68.7% similarity to Citrus and Poncirus pgip genes, respectively. Its open reading frame of the gene encodes a polypeptide of 330 amino acids, in which 10 leucine-rich repeats arrange tandemly. A new set of primers is designed to the 5′ and 3′ ends of the gene, which allows amplification of the full-length gene from the cotton cDNA library. Genomic DNA analysis reveals that this gene has no intron.     

黑线仓鼠MHCⅡ类DQA基因外显子2的克隆与序列分析

为了探明黑线仓鼠MHC的结构与功能并寻找分子标记,对MHCⅡ类DQA基因的外显子2进行克隆和序列分析．提取黑线仓鼠3个群体（吴村、沂南和临朐）的基因组DNA构建基因池,利用PCR技术扩增得到249bp的片段,将该目的片段连接到pMD18-T载体中,重组质粒转入大肠杆菌DH5α后利用蓝白斑法筛选阳性克隆,测序后得到该目的片段的核苷酸序列（Genbank登录号：FJ209306）并推导出氨基酸序列．结果表明：黑线仓鼠、人类、大鼠、小鼠、猪、马、牛、兔之间DQA基因外显子2的核苷酸序列同源性为68．7％-85％,氨基酸序列同源性为56．8％-83．5％,黑线仓鼠与大鼠、小鼠亲缘关系更近．测序得到的OQA基因外显子2的序列在物种间具有丰富的多态性,可以作为物种遗传分析的分子标记．     

油菜单株总DNA的快速制备

通过改进匀浆过程确定了适合同时提取多个１００ｍｇ左右植物材料总ＤＮＡ的实验程序,制备的总ＤＮＡ经测定紫外吸收曲线,限制酶作用和ＲＡＰＤ扩增,证明完全符合分子生物学实验的基本要求。     

紫外辐射诱导牡蛎细胞DNA损伤的单细胞凝胶电泳检测

以牡蛎为研究对象,建立了水产动物DNA损伤的单细胞凝胶电泳检测方法.分别以20W紫外灯距离30 cm照射牡蛎血液细胞30、60和120 s,然后用单细胞凝胶电泳技术检测细胞DNA的损伤.结果表明:未照射的细胞未受损伤,在电场中核DNA几乎不泳动,染色后呈圆形的荧光团,无拖尾现象.紫外照射后细胞核DNA均不同程度受损,且拖尾率、尾长、尾矩、Olive尾矩等指标值均随着照射时间加长而增加,DNA损伤程度与紫外照射时间之间存在明显的时间效应关系.研究结果为我国海洋环境中辐射防护研究和辐射毒理学检测提供新的研究方法和思路.     

Nucleotide sequence of the gene coding for the major protein of hepatitis B virus surface antigen.

DNA extracted from hepatitis B virus Dane particles has been cloned in bacteria using a plasmid vector. A full-length clone has been examined by restriction endonuclease analysis, and the nucleotide sequence of an 892-base pair fragment from cloned hepatitis B viral DNA encoding the surface antigen gene is reported. The amino acid sequence deduced from the DNA indicates that the surface antigens is a protein consisting of 226 amino acids and with a molecular weight of 25,398. The portion of the gene coding for this protein apparently contains no intervening sequences.     

Synthesis and pH Sensitive Fluorescent Performance of 2'-Chloroseminaphthofluorescein with Dual Bands

A novel unsymmetrical fluorescein derivative substituted by chlorine and naphthol, 2 '-chloroseminaphthofluorescein, was prepared by the two-step synthetic method. This method can effectively control the reactive process,so the pure unsymmetrical fluorescein derivative can be obtained in high yield. The new fluorophore exhibited dual absorption bands,dual excitation bands and dual emission bands with long-wavelength,selectively exciting the acidic and basic forms of the fluorophore. Compared with the similar symmetrical fluorescein derivatives such as 2 ', 7 '-dichlorofluorescein and dinaphtofluorescein, this fluorophore was found to have larger Stokes shift,especially in the basic form. Based on the properties above,this unsymmetrical fluorescein derivative is expected as a good fluorescent probe to be used where the interference exists due to the presence of endogenous fluorescers.