DNA生物传感器及其研究进展

就ＤＮＡ生物传感器的工作原理,分类、ＤＮＡ探针的固化方法,以及电化学ＤＮＡ生物传感器、光学ＤＮＡ生物传感器及压电ＤＮＡ生物传感器的研究进展、优缺点和发展趋势加以介绍。     

一种从重组质粒快速提纯DNA插入片段的方法

本文介绍一种从重组质粒中快速提纯ＤＮＡ插入片段的方法。质粒ＤＮＡ的制备简单、快速、分离的质粒ＤＮＡ可用于限制性酶切和转化大肠杆菌等。从琼脂糖凝胶中提纯ＤＮＡ插入片段的方法操作简单,回收效率高,提纯的ＤＮＡ片段可用于连接和制备杂交探针等。     

DNA芯片制作原理及其杂交信号检测方法

文章讨论了ＤＮＡ芯片的制作原理和杂交信号的检测方法。依其结构,ＤＮＡ芯片可分为两种形式,ＤＮＡ阵列和寡核苷酸微芯片。ＤＮＡ芯片的制作方法主要有光导原位合成法和自动化点样法。ＤＮＡ芯片与标记的探针或ＤＮＡ样品杂交,并通过探测杂交信号谱型业实现ＤＮＡ序列或基因表达的分析。适应于ＤＮＡ芯片的发展,同时出现了许多新型的杂交信号检测方法。主要有激光荧光扫描显微镜、激光扫描共焦显微镜、结合作用ＣＣＤ相机的荧光     

同时抽提DNA和RNA的简便方法

同时抽提ＤＮＡ和ＲＮＡ的简便方法刘定干（中国科学院上海生物化学研究所,２０００３１）关键词ＤＮＡＲＮＡ抽提在研究工作中,常同时需要培养细胞的ＤＮＡ和ＲＮＡ,从同一样品中同时提取ＤＮＡ和ＲＮＡ。目前,已有好几种同时抽提ＤＮＡ和ＲＮＡ的方法［１～４］。作...     

新书介绍

《ＤＮＡ 蛋白质相互作用》(ＤＮＡ ＰｒｏｔｅｉｎＩｎｔｅｒａｃｔｉｏｎｓ)ＡｎｄｒｅｗＴｒａｖｅｒｓ和ｍａｌｃｏｌｍＢｕｃｋｌｅ编著 ,2 0 0 0年ＯｘｆｏｒｄＵｎｉｖｅｒｓｉｔｙＰｒｅｓｓ出版 ,375页。蛋白质与ＤＮＡ的相互作用在分子生物学领域占有重要地位。例如 ,在确定和调节染色体结构、ＤＮＡ复制、重组和修复、基因转录、病毒感染等方面具有很大作用。本书是研究蛋白质与ＤＮＡ相互之间特异作用及许多研究方法的概述。描述了目前适用的许多主要的技术方法 ,尤其是分析转录及它与染色质结构关系的生物物理方法。每一种方法…     

一种提高质粒DNA凝胶电泳检测灵敏度的方法

比较了凝胶电泳示检测质粒ＤＮＡ时不同激发波长对ＤＮＡ－ＥＢ荧光强度的影响,发现短波长激发光可增加ＤＮＡ的探测灵敏度。采用２６０ｎｍ作为激光光时可探测到少至０．７ｎｇ的线性ＤＮＡ。且在很广的ＤＮＡ的质量范围内,ＤＮＡ－ＥＢ荧光强度 与ＤＮＡ量或正比。以此改进方法检测电离辐射诱导的ＤＮＡ单、双链断裂岢得到与其它研究结果相一致的Ｇ（ＳＳＢ）和Ｇ（ＤＳＢ）值。     

DNA分子标记技术及其在植物遗传多样性研究中的应用

本文阐述了ＤＮＡ限制性片段长度多态性、ＤＮＡ指纹图谱、单位点小卫星、单位点微卫星及随机扩增多态ＤＮＡ等主要的ＤＮＡ遗传标记技术的基本原理和方法．综述了不同ＤＮＡ标记的优缺点及其在植物遗传多样性研究中的应用前景．     

高效的植物DNA提取方法

利用液氮研磨植物幼芽,以苯酚─氯仿─异戊醇─核糖核酸酶法提取了大豆、菜豆、玉米、高梁等几种农作物的总ＤＮＡ。所提取的ＤＮＡ经岛津ＵＶ－２６５紫外分光光度计检测及０．６％的琼脂糖凝胶电泳,结果证明：该方法所提取的植物总ＤＮＡ纯度高,片段长度整齐,约５０ｋｂ左右,符合外源ＤＮＡ导入作物的要求,并且ＤＮＡ收率很高。     

分子间三链DNA的应用

自Ｄｅｒｖａｎ等１９８７年首先证实三链ＤＮＡ的形成可介导对靶ＤＮＡ的特异切割以来,有关三链ＤＮＡ的研究进展和奶快。本综述了近几年分子间三链ＤＮＡ在基因表达调控、染色体作图、基因分离及定点诱变等方面的应用,并对三链ＤＮＡ应用中存在的几个问题及相应的解决方法作了简单介绍。     

双歧杆菌总DNA对小鼠IL—2,NK活性影响的研究

目的 从双歧杆菌、大肠杆菌提取ＤＮＡ,用ＤＮＡ免疫小鼠,观察免疫功能的变化,探讨双歧杆菌ＤＮＡ对小鼠免疫功能的影响,并作对比研究。方法 肌肉注射提取的双歧杆菌ＤＮＡ、大肠杆菌ＤＮＡ,颈椎处死后,检测脾细胞的免疫功能,同时提取ＩＥＬ细胞与ＤＮＡ共孵育,检测它对ＩＥＬ细胞的激活情况及细胞因子产生情况,以自然杀伤细胞（ＮＫ）活性,白细胞介素２（ＩＬ－２）产生能力为指标,测定小鼠上述各项指标变化。结果 双歧杆菌ＤＮＡ、大肠杆菌ＤＮＡ肌肉注射后,小鼠以上两项指标与相应对照组相比较均明显提高（Ｐ〈０．０５）。双歧杆菌ＤＮＡ提高小鼠ＮＫ活性与ＩＬ－２水平程度大于大肠杆菌ＤＮＡ的作用（Ｐ〈０．０１）。结论 双歧杆菌ＤＮＡ可快速激活ＮＫ活性,提高体内ＩＬ－２水平。其效能优于大肠杆菌ＤＮＡ。     

Universal and rapid salt-extraction of high quality genomic DNA for PCR-based techniques.

A very simple, fast, universally applicable and reproducible method to extract high quality megabase genomic DNA from different organisms is described. We applied the same method to extract high quality complex genomic DNA from different tissues (wheat, barley, potato, beans, pear and almond leaves as well as fungi, insects and shrimps' fresh tissue) without any modification. The method does not require expensive and environmentally hazardous reagents and equipment. It can be performed even in low technology laboratories. The amount of tissue required by this method is approximately 50-100 mg. The quantity and the quality of the DNA extracted by this method is high enough to perform hundreds of PCR-based reactions and also to be used in other DNA manipulation techniques such as restriction digestion, Southern blot and cloning.     

改良CTAB法用于多年生植物组织基因组DNA的大量提取*

根据多年生植物组织富含多酚、多糖的具体特性,对现有的DNA提取方法进行了改进。通过增加提取缓冲液中b-巯基乙醇用量,简化氯仿/异戊醇抽提液步骤,改用经-20℃预冷异丙醇沉淀DNA等,对CTAB法加以改进。改进后方法具有以下优点：（1）获得的DNA质量良好,提取过程无明显的DNA降解,基本上排除了多酚物质的干扰;（2）用获得的DNA进行Southern杂交,可得到理想的杂交信号,可满足相关的分子研究要求;（3）操作简便。Abstract It is a difficult problem to isolate high quality DNA from plants containing a high contents of polyphenolics and polysaccharose, such as Actinidia plant. The protocol described in this paper is a modified CTAB (hexadecyltrimethylammonium bromide) method. High quality genomic DNA can be isolated from Actinidia plant using the improved method. The DNA is good enough for Southern blot and other uses in DNA research. The protocol is also efficient for quick and macro-DNA extraction.     

一种改良的植物基因组DNA通用提取方法

高质量的基因组DNA是分子生物学研究的基础,而从富含糖类和次生代谢物且异质性强的植物材料中分离DNA相对困难。本方法在CTAB法和商业DNA提取试剂盒的基础上,在裂解细胞之前,对植物材料进行预处理．去除干扰DNA提取的代谢物,并在后续步骤中进行了一些优化。该方法适于多种不同的植物种类,所提取的基因组DNA质量较好,能满足下一步基因操作的要求,是一种通用的植物基因组DNA提取方法。     

一种高效的哺乳动物粪便DNA提取通用方法

以粪便为材料提取动物DNA进行动物保护遗传学和分子生态学研究的关键是能否提取到高质量的粪便DNA.然而提取方法通用性不好和产物质量不高等问题阻碍了粪便DNA分析技术的推广.本文介绍的改进型十六烷基三甲基溴化铵提取法可广泛适用于各食性哺乳动物粪便DNA提取,在11种不同食性动物的粪便DNA提取实验中验证了它的可靠性和通用性.本方法成本低廉(3元/样),用实验室常规试剂即可完成粪便DNA提取,其产物纯度高于专用试剂盒QIAamp DNA Stool Kit,在拥有超过专业试剂盒提取效果的同时尽可能的降低了实验成本,有利于粪便DNA技术的推广.     

苔藓植物DNA提取方法研究

提取高质量的 DNA是对苔藓植物遗传多样性进行研究的基础。该文以苔藓植物为试材 ,用 5种方法 ,即快速提取法、改良 CTAB法、CTAB法、SDS法及高盐法 (第一种为自行设计 ,第二种是对原有方法的改进 )对苔藓植物 DNA提取方法进行了比较研究。结果表明 ,快速提取法和改良 CTAB法是 2种适合于苔藓植物 DNA提取的方法。这 2种方法提取的 DNA浓度和纯度均比较高 ,凝胶电泳显示无明显降解现象 ,适宜作为 PCR扩增的模板 ,并成功地进行了 RAPD扩增。     

A pressure cooking-based DNA extraction from archival formalin-fixed, paraffin-embedded tissue

As emerging novel DNA-based methodologies are adopted, nucleic acid-based assays depend critically on the quality and quantity of extracted DNA. Formalin-fixed, paraffin embedded (FFPE) tissue samples provide an invaluable resource for subsequent molecular studies of clinical phenotypes, but high-quality DNA extraction from archival FFPE tissue specimens remains complex and time-consuming. To address this challenge, we have developed a reliable rapid DNA extraction method for FFPE tissue specimens. It is based on deparaffinization at high temperature coupled with relieving crosslink in a pressure cooker. The DNA yield by this rapid method resulted in an average 1.8-fold increase in comparison with the commercial kit and OD 260/280 ratios between 1.87 and 1.95. The DNA obtained by the rapid method was suitable for methylation analyses in colon cancer patients. These data suggest that this new DNA extraction method coupled with methylation-specific polymerase chain reaction can be used for epigenetic studies with the advantages of rapidity and high quality and may contribute to the development of biomarkers in clinical studies.     

改进的SDS-CTAB法提取濒危植物连香树总DNA

对珍稀濒危植物连香树(Cercidiphyllum japonicum)的6种总DNA提取方法进行了对比试验,结果表明改进的SDS-CTAB法更适合于连香树总DNA提取。该方法提取的DNA经紫外消光值检测,其A260/A280为1.8532,优于CTAB法(1.4872)、SDS法(1.3552)、PVP法(1.5079)、尿素法(1.1858)和高盐低pH法(1.4534)。琼脂糖凝胶电泳和PCR扩增结果也得出同样的结论。     

Evaluation of mailed pediatric buccal cytobrushes for use in a case-control study of birth defects

BACKGROUND: Buccal cell collection is a convenient DNA collection method; however, little attention has been given to the quality of DNA obtained from pediatric populations. The purpose of this study was to determine the effect of a modified cytobrush collection method on the yield and quality of infant buccal DNA collected as part of a population-based case-control study of birth defects. METHODS Cytobrushes were collected from infants, mothers, and fathers using a standard collection method in 1997 to 2003 and a modified protocol that allows air-drying of the cytobrushes after collection from 2003 to the present. Yield and quality of DNA from 1057 cytobrushes was assessed by quantitative PCR and short tandem repeat (STR) genotyping, respectively. RESULTS Air-dried cytobrushes from infants had higher median DNA yields (1300 ng) and STR completion rates (99.5%) than standard collection method cytobrushes (60 ng and 59.5%, respectively). A subset of DNA aliquots was genotyped for six single nucleotide polymorphisms (SNPs). Aliquots from both collection methods that passed the quality protocol (DNA concentration >1 ng/μl, and successful amplification of ≥1 STR) had high genotype completion rates (99-100%). The median DNA yield following whole genome amplification was more than twofold higher for air-dried than standard collection specimens (p < 0.001). CONCLUSION Yield and quality of buccal DNA collected from infants are improved by using a method that incorporates air-drying; however, DNA collected by both methods is suitable for genotyping if stringent quality control procedures are instituted. These findings may be helpful for future epidemiologic studies of birth defects and other adverse pediatric outcomes.     

一种直接用于PCR扩增的山羊瘤胃微生物DNA提取方法的建立

目的建立提取高质量的瘤胃微生物DNA的方法,为采用免培养技术研究山羊瘤胃微生物奠定基础。方法采集山羊瘤胃内容物,用SDS高盐法提取微生物总DNA,以通用引物扩增细菌和古细菌的16SrDNA。结果提取到的瘤胃微生物总DNA片段大于23kb,PCR能够扩增出细菌和古细菌的16SrDNA片段。结论用该提取方法得到的山羊瘤胃微生物总DNA能够满足后续实验的需要。     

An improved method of plant megabase DNA isolation in agarose microbeads suitable for physical mapping and YAC cloning

The isolation of high quality megabase DNA from plant cells that is susceptible to a variety of molecular reagents is a critical first step in the physical analysis of complex genomes. A method for the isolation of such DNA by encapsulating plant protoplasts in agarose microbeads is presented. In comparison with the conventional agarose plug method, microbeads provide a dramatic increase in the surface area yielding megabase DNA that can be treated essentially as an aqueous DNA solution. Examples of the utility of DNA prepared by this technique for physical mapping, partial restriction enzyme digestion and cloning of large inserts as YACs are presented.