DNA链置换技术的研究现状与展望

综述了利用DNA分子元件构造人工逻辑生化电路的新技术——DNA链置换技术在构造逻辑门运算模型、生化逻辑电路与神经网络、DNA纳米机器人、DNA反应网络等领域的研究进展,并通过对半加器/全加器逻辑运算模型和编码器逻辑运算模型的设计及仿真,对DNA链置换技术的相关应用进行了实验验证.在此基础上提出:构建运动及功能型DNA纳米机器,整合DNA逻辑门、自底向上地构建DNA计算机体系结构,将是DNA链置换技术应用的发展方向.     

汉族传统男服的造型及装饰DNA

立足于汉族传统服饰精粹的准确传承与发展,以历史学、社会学、设计学等理论为指导,通过文献调研、实地考察等对以中原为核心地域的汉族传统男服造型与装饰之典型特征进行了系统研究。在对其概念、地域环境研究的基础上,归纳了造型与装饰DNA体系,具体包括造型之廓形DNA、结构DNA、材质DNA、技艺DNA,装饰之元素DNA、形式DNA、色彩DNA、质料DNA、工艺DNA。研究认为,古汉族男服是规范社会秩序、服务政治伦理的工具,其DNA体系彰显了"合乎天道"的服饰审美情趣与设计规则。     

食物成分对肠道细胞DNA双链断裂损伤的保护作用及机制研究进展

DNA双链断裂作为最严重的DNA损伤是引起人体基因组不稳定和引起基因疾病形成的潜在因素。研究发现,多种食物成分具有抑制DNA双链断裂、促进DNA损伤修复的功能。本文主要概括了近20 年来国内外报道的对DNA双链断裂损伤具有调节功能的食物成分,从抗氧化作用、调节DNA损伤修复和抗氧化同时调节DNA损伤修复3 个方面阐述了食物成分缓解肠道细胞DNA双链断裂损伤的作用机制,并探讨了不同摄入时间和摄入剂量对DNA双链断裂损伤保护效果的影响,对可能缓解DNA双链断裂损伤的其他潜在食物成分进行展望,为开发新型缓解DNA双链断裂损伤的食物提供理论参考。     

葡萄酒中葡萄的DNA提取和分子鉴定研究进展

分子生物学技术能够对葡萄酒真伪和品质鉴别提供直接的判别依据。解决葡萄酒中葡萄DNA提取的技术难题,并以此为基础利用分子标记技术进行葡萄酒中葡萄品种的鉴定,具有重大的研究和应用价值。因此,本文综述了近10余年来分子生物学技术在葡萄酒分子鉴定方面的研究进展,从葡萄酒中葡萄DNA提取、DNA质量评价和品种鉴定三方面进行了阐述,尤其是将文献中的DNA提取和下游检测技术方案进行了系统比较,就DNA提取部分按照葡萄DNA的富集、DNA粗提和多糖多酚的去除以及DNA纯化回收3 个步骤进行了深入的分析,并详细介绍了以微卫星标记为代表的品种鉴定技术现状。在此基础上,本文还探讨了基因检测技术在葡萄酒真伪和品质鉴定方面的应用前景。     

GA118系列遗传分析仪数据采集软件的设计和实现

GA118系列法医DNA专用检测平台包括GA118系列法医DNA遗传分析仪、法医DNA遗传分析仪配套耗材和数据采集软件。主要应用于法医DNA鉴定实验室,实现PCR产物的基因分型。用户通过此平台中的数据采集软件控制法医DNA遗传分析仪,实现DNA荧光光谱数据采集,生成通用数据格式的DNA数据文件,提供给后续的DNA分析软件使用。文章从整体的角度介绍了GA118系列数据采集软件,包括软件的总体设计思路、软件架构设计、软件接口设计、软硬件之间的通讯及控制方式、软件中各模块实现的功能说明等。     

茶籽油DNA提取方法的比较

针对食用油脂中DNA含量极低、DNA序列片段短、破坏严重的特点,通过实时荧光PCR法检测植物的通用tRNALeu,对茶籽油DNA几种提取方法进行比较,建立了食用油脂中DNA提取新方法。结果证明:较之于文献中常见植物油DNA几种提取方法,用改进方法提取的DNA含量高、纯度高,可以作为PCR反应的模板,为食用油脂进行核酸类生物性检测提供了一种简捷有效的方法。     

银离子掺杂DNA的拉曼光谱研究

文章研究了掺杂了银离子的DNA的拉曼光谱。结果显示结合了银离子的DNA发生了一系列的变化,鸟嘌呤和腺嘌呤的伸缩振动峰1576、1487、1375和727cm-1向低波数移动了4-10cm-1,碱基之间氢键的拉曼谱峰1665cm-1明显变宽,DNA中PO2-的对称伸缩振动峰1091和788cm-1分别移动到了1085和781cm-1,B构象830cm-1谱峰强度明显降低。说明银离子和DNA碱基之间的氢键产生了相互作用,使DNA双链解旋,并且DNA的构象也发生改变。     

梯度金黄色葡萄球菌DNA杂交分子信标激发荧光初探

验证不同浓度黄色葡萄球菌DNA与特殊设计的分子信标探针杂交,观察荧光强度的变化,并验证该探针对于金黄色葡萄球菌DNA的特异性。利用特殊ATMND染料特异性嵌入缺失DNA位点的独特分子信标设计,并巧妙利用了能与DNA茎环结构杂交互补金黄色葡萄球菌特异DNA序列,导致茎环结构构相改变,而使ATMND染料离开缺失位点被重新激发荧光。将不同浓度黄色葡萄球菌DNA与分子信标探针杂交,观察荧光强度的变化,并选择标准菌株DNA对该探针进行特异性验证。证实了这种探针能够特异性的选择杂交金黄色葡萄球菌目标序列,随着金黄色葡萄球菌DNA浓度的增强,荧光信号也随之增强,与其它沙门氏菌、大肠杆菌、阪崎肠杆菌、单核细胞增生李斯特菌和铜绿假单胞菌DNA相比,具有更为明显的荧光恢复功能。荧光信号随着金黄色葡萄球菌DNA浓度的增高而增强,该探针在金黄色葡萄球菌DNA的识别上具有一定的特异性。     

应用UV-X-IP分析酿酒酵母HSF和靶DNA的相互作用

利用紫外激光交联和免疫沉淀技术(UV laser crosslinking and immunoprecipitation, UV-X-IP)分析原核表达的酿酒酵母热激因子(heat shock factors,HSFs)和片段化的基因组靶DNA的相互作用.分别在ScSSA1、ScSSA4启动子区和编码区设计引物对免疫沉淀的DNA 进行PCR分析.结果发现,紫外激光交联和免疫沉淀DNA库中富集了ScSSA1、ScSSA4启动子区HSF结合的DNA片段,而没有编码区DNA,表明HSF特异结合了靶DNA.从未经紫外激光交联样品的免疫沉淀DNA库中未获得相应PCR扩增产物,说明紫外激光在酿酒酵母HSF与靶DNA之间形成共价结合中起重要作用.     

酵母核酸常用的鉴定方法及其应用

随着酵母全基因组测序的完成,针对酵母的染色体、基因组DNA、核糖体RNA(r RNA)和线粒体DNA(mt DNA)采用分子生物学技术手段进行已知酵母菌的鉴定,并用于未知菌株多样性分析的方法得到了越来越多的应用。综述并对比了酵母菌的分子生物学分类鉴定方法及其实际应用,其中包括PFGE、r RNA基因的18S、26S,5.8S-ITS区域的PCR、PCR-RFLP、PCR-单链构象多态性分析、DNA的RAPD分析、线粒体DNA的酶切分析等。     

Comparison of four commercial DNA extraction kits for PCR detection of Listeria monocytogenes, Salmonella, Escherichia coli O157:H7, and Staphylococcus aureus in fresh, minimally processed vegetables

Four commercial DNA extraction methods, PrepMan Ultra (Applied Biosystems), InstaGene Matrix (BioRad), DNeasy Tissue kit (Qiagen), and UltraClean (MoBio), were tested for PCR detection of Listeria monocytogenes, Escherichia coli O157: H7, Salmonella, and Staphylococcus aureus in fresh, minimally processed vegetables. For comparative purposes, sensitivity assays with specific PCRs were carried out after DNA extraction with the four methods in green pepper, broccoli, and onion artificially inoculated with the four pathogens separately. As confirmed by statistical analysis, the DNeasy Tissue kit rendered the highest sensitivity values in the three matrices assayed for Salmonella, L. monocytogenes, and E. coli O157:H7 and in onion for S. aureus. Despite being the most expensive of the methods compared, the DNeasy Tissue Kit can be successfully applied for any of the four most commonly studied pathogens, thus saving time and overall reducing the cost of the analysis.     

浓香型白酒黄水、窖泥中微生物总DNA提取方法比较

为高效地获得浓香型白酒窖池中窖泥和黄水中的微生物总DNA以更好地分析其过程中微生物区系,该文设计了4种不同的总DNA提取方法,分别提取了泸州某知名浓香型白酒企业黄水和窖泥的DNA,并用紫外吸收和PCR-DGGE方法比较了不同提取方法所获得总DNA的纯度及其所代表的原核微生物多样性状况。结果显示,4种提取方法均能从2种样品中提取到纯度较高的DNA;以这些DNA为模板均能扩增出细菌和古菌16S rDNA的部分片段的特异性条带;其DGGE电泳图谱均呈现出较为丰富的条带多样性,但条带存在明显差异。综合评价DNA纯度、扩增效果以及DGGE谱图,对于黄水和窖泥,“酶+SDS+液氮法”提取样品中原核微生物DNA的效果最优。     

PCR 检测牛乳中大肠杆菌基因组DNA 的提取方法

为获得一种适于牛乳样品大肠杆菌PCR 检测的基因组DNA 提取方法,对饱和酚法、CTAB 法、试剂盒法及溶剂裂解法等4 种提取方法加以比较,通过考察DNA 的纯度、定量分析以及PCR 分析,确定一套有效、快速、适合牛乳中提取大肠杆菌的基因组DNA 方法--溶剂热裂解法。结果显示该方法提取的DNA 模板,适于PCR扩增大肠杆菌DNA,可以用于牛乳中的大肠杆菌检测。     

Polymerase chain reaction-mediated characterization of molds belonging to the Aspergillus flavus group and detection of Aspergillus parasiticus in peanut kernels by a multiplex polymerase chain reaction

The Aspergillus flavus group covers species of A. flavus and Aspergillus parasiticus as aflatoxin producers and Aspergillus oryzae and Aspergillus sojae as koji molds. Genetic similarity among these species is high, and aflatoxin production of a culture may be affected by cultivation conditions and substrate composition. Therefore, a polymerase chain reaction (PCR)-mediated method of detecting the aflatoxin-synthesizing genes to indicate the degree of risk a genotype has of being a phenotypic producer was demonstrated. In this study, 19 strains of the A. flavus group, including A. flavus, A. parasiticus, A. oryzae, A. sojae, and one Aspergillus niger, were subjected to PCR testing in an attempt to detect four genes, encoding for norsolorinic acid reductase (nor-1), versicolorin A dehydrogenase (ver-1), sterigmatocystin O-methyltransferase (omt-1), and a regulatory protein (apa-2), involved in aflatoxin biosynthesis. Concurrently, the strains were cultivated in yeast-malt (YM) broth for aflatoxin detection. Fifteen strains were shown to possess the four target DNA fragments. With regard to aflatoxigenicity, all seven aflatoxigenic strains possessed the four DNA fragments, and five strains bearing less than the four DNA fragments did not produce aflatoxin. When peanut kernels were artificially contaminated with A. parasiticus and A. niger for 7 days, the contaminant DNA was extractable from a piece of cotyledon (ca. 100 mg), and when subjected to multiplex PCR testing using the four pairs of primers coding for the above genes, they were successfully detected. The target DNA fragments were detected in the kernels infected with A. parasiticus, and none was detected in the sound (uninoculated) kernels or in the kernels infected with A. niger.     

冷却猪肉不同前处理对细菌DNA提取及PCR-DGGE的影响

比较研究冷却猪肉不同前处理对细菌DNA 提取及PCR- 变性梯度凝胶电泳(DGGE)的影响。冷却猪肉4℃贮藏至第5 天时,分别采用冻融、超声、摇床和匀浆前处理后,提取细菌DNA,进行Dilution-PCR 和DGGE 分析。结果表明,不同前处理方法所制备的DNA 量稍有差别,但对PCR-DGGE 结果无显著影响,故可根据实际情况选择上述不同前处理方法。同时对16S rDNA V3 区的DGGE 图谱进行割胶测序,检测到腐败菌主要是假单胞菌属,其他还有不动杆菌属、环丝菌属和沙雷氏菌属。     

Genetic procedures for identification of enterotoxigenic strains of Staphylococcus aureus from three food poisoning outbreaks

Three food poisoning restaurant outbreaks due to Staphylococcus aureus, occurring during June-October 2002 in the Principality of Asturias (PA), Spain, provided the basis for investigating some aspects of the molecular epidemiology of this organism. The methods applied to identify strains and lineages included multiplex-polymerase chain reaction (PCR) to detect nine enterotoxin (se) genes, and three DNA fingerprinting procedures: pulsed-field gel electrophoresis (PFGE) with SmaI, randomly amplified polymorphic DNA (RAPD) with two selected primers, and plasmid restriction analysis with HindIII. Thirty-two isolates were differentiated into three non-se and 12 se strains, which were outbreak-specific, except for one that was represented in two of the outbreaks. In outbreak 1, the 16 food isolates analyzed had sec, seg and sei genes and generated a distinctive DNA fingerprint, being assigned to a single strain. This strain could be categorized as endemic in the PA and associated to manually handled dairy products and nasal carriers. In outbreak 2, the four food isolates analyzed fell into three strains, each one displaying a different se-gene profile (sea, sec and seg-seh-sei) and a distinctive DNA fingerprint. In outbreak 3, the five food isolates tested fell into four seg-sei strains generating identical RAPD but different PFGE and plasmid profiles, and one sea strain also collected from two nasal carriers. This last strain had also been found in manually handled vegetables in outbreak 2, and it belongs to a not very frequently found sea lineage in the PA. Multiplex-PCR to detect se genes together with the three applied DNA fingerprint typing procedures proved therefore to be useful tools in subclassifying S. aureus for epidemiological purposes.     

普洱茶发酵过程中微生物总DNA提取方法的比较

为了快速提取普洱茶发酵过程中微生物总DNA,便于分析微生物群落的多样性及演替情况,对比研究了总DNA提取的四种方法-酶法、试剂盒法、超声波法和变性剂法,以16S rRNA区域通用引物(27F和1492R)和18S rRNA区域通用引物(NS1和NS8)对四种方法提取的总DNA分别进行扩增,根据总DNA提取的大小、产量、纯度和PCR扩增产物进行评估。综合各项指标来看,总DNA提取方法以变性剂法为优,其次是试剂盒法、超声波法和酶法。     

Lyophilisation improves the extraction of PCR‐quality community DNA from pig faecal samples

BACKGROUND: Faeces are increasingly used as sources of DNA for genetic and ecological studies. Although multiple methods to preserve faecal samples prior to DNA extraction have been used (e.g., 70% or absolute ethanol, freezing at ?20 °C or in liquid nitrogen) no information is at present available in the literature on the use of lyophilised faeces. Accordingly, the yield and quality of the community DNA obtained by using four different commercial DNA extraction kits (QIAamp DNA Stool Mini Kit, REALPURE Spin Kit, SPEEDTOOLS Tissue DNA Extraction Kit, and JETQUICK Tissue DNA Spin Kit) from fresh and lyophilised samples of faeces were studied here. RESULTS: The use of lyophilised faeces resulted in a 1.5‐ to 2‐fold increase in DNA recovery relative to the use of fresh faeces regardless of the kit used. Among the four kits tested, the best results were obtained with the QIAamp DNA Stool Mini Kit. Community DNA obtained from lyophilised faeces also provided the best restriction fragment length polymorphism (PCR–RFLP) profiles, which should guarantee a better representation of the microbial diversity present in faecal samples. CONCLUSION: As compared with using fresh faecal samples for pig faecal microbiota studies, lyophilisation improved both DNA yield and quality of the information arising from the PCR–RFLP method of analysis. Copyright © 2009 Society of Chemical Industry     

食品检测用4种病原菌基因组试剂盒提取方法的比较与优化

目的:采用国产的天根细菌基因组提取试剂盒,制备具有自主知识产权的、达到国家核酸标准物质要求的食源性病原菌的基因组样品。方法:以4种食源性病原菌大肠埃希氏菌O157:H7、单增李斯特氏菌、肠炎沙门氏菌和金黄色葡萄球菌为研究对象,首先使用进口OMEGA细菌基因组提取试剂盒和国产天根细菌基因组提取试剂盒提取4种病原菌的基因组,然后对国产试剂盒提取的基因组浓度和纯度较低的大肠埃希氏菌O157:H7、肠炎沙门氏菌和金黄色葡萄球菌的基因组提取方法进行优化。最后对4种病原菌基因组的完整性和纯度进行琼脂糖凝胶电泳检测,并分别对4种病原菌的特征毒力基因进行PCR扩增验证。结果:通过方法优化,使用国产天根细菌基因组提取试剂盒提取的4种病原菌的基因组质量浓度均在50 ng/μL以上,A260/A280和A260/A230的值在1.7~2.0之间,基因组DNA条带完整,无RNA和蛋白污染,各菌株的特征毒力基因均为阳性。结论:通过基因组提取方法的优化,得到符合国家核酸标准样品要求的4种病原菌基因组样品,为后续大量制备核酸标准样品奠定了基础。