一步法提取植物DNA用于大规模RAPD分析

RAPD技术已广泛地用于植物的系统演化、种群多样性、群体遗传学及杂种种子纯度的鉴定等工作中[1～3].本文基于快速节省的原则对碱液法提取植物DNA进行一些改进.     

适于RAPD分析的真菌DNA提取方法

报道一种高纯度、高分子量真菌基因组DNA的快速小量提取方法。该法制备的DNA降解少,分子量均在48.5kb以上;纯度高,所有样品的A260/280都在1.7-2.0之间;产率也很高,一般从500mg干菌丝可稳定获得530μg的DNA。所获得的DNA适用于RAPD分析。     

几种提取枣和酸枣DNA用于RAPD分析的方法比较

利用不同的方法提取枣和酸枣的总DNA,并分析了不同样品状况、抗氧化剂和纯化过程等对所提DNA质量及其RAPD扩增效果的影响。实验显示：用改良CTAB法优于SDS法,可有效去除多糖。加入的抗氧化剂PVP、抗坏血酸、β-巯基乙醇等可有效阻止多酚类物质褐化,但不同抗氧化剂间效果差别不大。分别将样品干燥处理、固定处理、液氮处理、冷藏,与新鲜材料相比,用液氮处理并保存于-80℃的材料与新鲜材料所提DNA相当,而用其他方法DNA都有不同程度降解。不同材料提取结果比较显示：幼叶所提DNA产量和质量优于老叶,并且不需要较多的纯化过程。在有RNA和少量蛋白质时,对扩增结果影响不大。     

栗属植物基因组DNA的提取及RAPD、SSR分析

以中国板栗、茅栗和锥栗的叶片为材料,提取栗属的DNA。针对栗属植物富含多酚类等次生物质的特点对栗属DNA的提取方法进行了改良,采用CTAB-蛋白酶K法加重复抽提,去除栗属植物中的相关次生物质,获得了高质量的DNA。所提取的DNA完全满足PCR、RAPD、SSR等一些分子生物学实验要求。     

供RAPD用高梁模板DNA的提取方法

植物总ＤＮＡ的提取方法一般多采用Ｄｏｙｌｅ等（１９９０年）的ＣＴＡＢ法进行操作。但由于试材不同及操作过程一些细节问题的处理不当，有时结果总是不尽人意，如断裂严重出现弥散及纯度不够等问题。本文以高粱为试材，在多次试验的基础上，摸索出一种适于ＲＡＰＤ用的...     

石斛干品基因组DNA的提取与RAPD分析

市场中药干品的药性差异一直是影响中药标准化的瓶颈,而检测技术相对落后是导致这一现象的主要原因。DNA分子水平检测的困难是药材干品的基因组DNA难以提取。本文以铁皮石斛(Dendrobium candidum)干茎为材料,采用了四种方法从干品石斛中提取基因组DNA。结果表明,采用改良的CTAB法可从石斛干品尤其是干茎皮中提取质量较高的基因组DNA,其分子量大于23kb,以此DNA为模板进行不同引物的PCR扩增可获得清晰的RAPD条带。该研究初步建立了石斛干品合适的RAPD技术体系。     

金弹总DNA提取及RAPD体系优化

以金弹叶片为材料,研究其总DNA提取方法及RAPD-PCR条件。结果表明,用改良CTAB法Ⅱ提取的DNA适于RAPD分析;优化的金弹RAPD-PCR体系为:反应体积20μl,Mg²⁺2.5 mmol/L、dNTP 0.25 mmol/L、引物0.20μmol/L、模板DNA 1.0 ng/μl和1 U Taq DNA聚合酶。相应的扩增程序为:94℃预变性3 min;94℃变性45 s,37℃复性60 s,72℃延伸120 s,循环45次;72℃延伸10 min,4℃结束。     

大血藤DNA提取及RAPD研究初探

以浙江天台山大血藤为材料,对其DNA提取和RAPD分子标记方法进行了研究。结果表明:所采用的经改进的SDS提取法可获得高质量的大血藤DNA,分子量大于23kb,可满足RPAD扩增;用15个不同的随机引物对所提取的12个个体的大血藤DNA进行了RAPD分子标记分析,其中14个引物扩增产物具有多态性。建立了大血藤DNA提取和RAPD标记的分析程序,为RAPD分析应用于大血藤遗传多样性研究打下了良好的基础。     

广东野百合DNA提取和RAPD条件的优化

以野百合(Lilium brownii)新鲜叶片、硅胶干燥叶片及鳞片为材料,研究了DNA的提取方法,并对影响随机扩增多态DNA(RAPD)反应的各因素进行了优化。建立了野百合RAPD的优化反应体系及程序,即在20μl反应体系中,含20 ng模板DNA,2.0 mmol/L Mg2 、0.2 mmol/L dNTPs、1.5 U Taq DNA聚合酶、0.3μmol/L随机引物S1519;扩增程序为:94℃预变性5 min,然后94℃30 s,38℃50 s,72℃1 min,35个循环,最后72℃延伸10 min,4℃保存。     

不同保存方式下蝗虫组织DNA的提取及RAPD分析

为了开展蝗虫分子系统学研究,分别对冷冻、乙醇浸泡(100%、乙醇、70％乙醇)和干制蝗虫标本用饱和NaCl法进行了基因组DNA的提取,并用随机引物进行扩增,结果表明：70％乙醇固定的标本和部分干标本提取的总DNA得率较低,在琼脂糖凝胶电泳检测中大音琏分有明显降解,导致PCR扩增中信息缺失,甚至无扩增条带;而保存完好的干标本、-20℃冷冻标本和100%,乙醇浸泡标本提取的总DNA带型整齐,无拖尾,PCR扩增结果的稳定性好,成为蝗虫分子系统学研究中首选的三种保存方式。     

Isolation of Pinus radiata Genomic DNA Suitable for RAPD Analysis

A protocol is presented for Pinus radiata genomic DNA isolation based on an alkyltrimethyl-ammonium bromide (CTAB) method described for other woody species. The method involves mortar grinding of tissue, a modified CTAB extraction employing high salt concentrations and polyvinyl pyrrolidone, a RNase A treatment and successive isoamyl alco- hol-chloroform extractions. The yield was approximately 15 g DNA per 100 mg of initial fresh plant material. The genomic DNA obtained by this method was suitable to be used in simple sequence repeat and random-amplified-polymorphic DNA reactions. This extraction method would allow the molecular analysis of shoots from different clones within P. radiata families.     

RAPD分析用的犁DNA提取方法

ＲＡＰＤ分析用的梨ＤＮＡ提取方法胡春根郝玉邓秀新史永忠（华中农业大学作物遗传改良国家重点实验室,武汉４３００７０）ＤＮＡＥｘｔｒａｃｔｉｏｎＭｅｔｈｏｄｆｏｒＲＡＰＤＡｎａｌｙｓｉｓｉｎＰｅａｒｓＨＵＣｈｕｎｇｅｎＨＡＯＹｕＤＥＮＧＸｉｕｘｉｎＳＨＩ...     

基于RAPD分析用百合DNA的提取

对百合DNA的提取过程进行改良和优化,筛选出较理想的DNA提取方法。结果表明,该方法提取的百合DNA在1％的琼脂糖胶电泳上为清晰的一条带,RNA去除干净,无降解现象,分子量大于23kb,OD200nm/OD280nm值为1．80-2．00。DNA的质量符合百合RAPD（随机扩增多态DNA）分析要求,在百合遗传多态性分析和品种分子鉴定中具有良好的应用前景．     

One Step Isolation of Bovine Asialoglycoprotein Receptor and Its Characterization by Sequence Analysis and MALDI Mass Spectrometry

The asialoglycoprotein receptor (ASGP-R), which is responsible for the uptake of partially deglycosylated serum glycoproteins was isolated from bovine liver. The receptor was purified in one step from solubilized plasma membranes by affinity chromatography on 6-(-D-lactosyl)-n-hexylamine coupled to N-hydroxysuccinimide activated Sepharose with a coupling degree of 7.6 mol/ml gel. The preparation yielded two distinct polypeptides with apparent molecular weights of 48 and 43 kDa as determined by sodium dodecyl sulfatepolyacrylamide gel electrophoresis. A polyclonal antibody raised against the human ASGP-R recognized the bovine 43 kDa protein in Western blot analysis. The 48 and 43 kDa polypeptides were digested by trypsin and the digests were subsequently analyzed by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Sequence analysis of four tryptic fragments, two each of the 48 kDa and of the 43 kDa polypeptides revealed that these were highly homologous to ASGP-R subunits from man, mouse and rat.     

一步法快速质粒鉴定方法

对《分子克隆实验指南》(第三版)中的牙签法小量制备质粒DNA的方法做了改进,减少了 加样次数,免去冰浴、离心等步骤,改进后的方法在琼脂糖凝胶电泳前仅需一步加样和加热过程, 提高了实验效率。同时结合使用多孔道移液器和96孔板,更适合于在高通量筛选中鉴定阳性克 隆。实验对改进后的方法与《分子克隆实验指南》上的方法进行了比较,表明“一步法”的检测效 果、准确率与重复性均有到较好的结果。     

The Isolation of DNA from Plant Materials

Total DNA was isolated from seedling and tissues of many plants. The nuclei and chloroplasts were prepared from plant tissues, and then the nuclear DNA and chloroplast DNA were isolated from them. According to the chemical analysis and the physical properties determined by ultraviolet absorbance, hyperchromicities, ultracentrifugation, gel electrophoresis and the electro-microscopical observations it is suggested that DNA obtained possessed a considerable purity and to a certain extent retained the natural status of the large molecule.     

濒危植物浮叶慈姑遗传多样性的RAPD分析

采用RAPD分子标记技术,对内蒙古二卡河(EK)和黑龙江库都尔(KDE)的浮叶慈姑居群进行了遗传多样性分析。 从60个随机引物中筛选出14个引物对2个居群共48个样品进行扩增,得到93条清晰的条带,其中70条具有多态性,即物 种水平上的多态位点百分率(PPB)为75.27%。POPGENE分析结果表明:二卡河居群的PPB为66.67%,库都尔居群的PPB 为72.04%。两个居群间基因分化系数(Gst)为0.0628,即遗传变异有很大一部分是来自居群内(93.72%)。结合相关的分析 表明生态学因素可能是浮叶慈姑濒危的主要原因。     

One Giant Leap for Categorizers: One Small Step for Categorization Theory

We explore humans’ rule-based category learning using analytic approaches that highlight their psychological transitions during learning. These approaches confirm that humans show qualitatively sudden psychological transitions during rule learning. These transitions contribute to the theoretical literature contrasting single vs. multiple category-learning systems, because they seem to reveal a distinctive learning process of explicit rule discovery. A complete psychology of categorization must describe this learning process, too. Yet extensive formal-modeling analyses confirm that a wide range of current (gradient-descent) models cannot reproduce these transitions, including influential rule-based models (e.g., COVIS) and exemplar models (e.g., ALCOVE). It is an important theoretical conclusion that existing models cannot explain humans’ rule-based category learning. The problem these models have is the incremental algorithm by which learning is simulated. Humans descend no gradient in rule-based tasks. Very different formal-modeling systems will be required to explain humans’ psychology in these tasks. An important next step will be to build a new generation of models that can do so.