共查询到20条相似文献,搜索用时 335 毫秒
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野生芋属植物干叶片DNA的提取及PCR扩增 总被引:1,自引:0,他引:1
野生芋属植物体内多糖、色素、酚类等次生物质含量较高,严重影响从中提取的DNA的质量.针对这一问题,作者以6种芋属植物的干叶片为材料,摸索出一种适合芋属植物的DNA提取方法,并对提取的DNA进行了纯度鉴定和PCR检测,结果表明此方法可有效去除次生物质对DNA的干扰,样品DNA的质量和纯度较高,可用于下游分子生物学操作. 相似文献
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顽拗植物类群的总DNA制备 总被引:43,自引:0,他引:43
从富含多糖的顽拗植物类群提取与纯化DNA是许多研究领域例如居群生物学、生物多样性、分子标记辅助育种研究普遍遇到的难题。以西南桦 (Betulaalnoides)为例发展了一套改进的方案 ,有效地从这种顽拗植物的干叶和鲜叶中制备了DNA。此方案包括 3个关键步骤 :首先从植物细胞匀浆中用不含CTAB的缓冲液洗去大部分多糖和其他次生物质 ;在提取介质中采用 3%CTAB而不是通常用的 2 %CTAB ;将常用的高盐去糖的纯化操作提前到用异丙醇沉淀DNA之前进行。从西南桦提取的DNA已成功地用于RAPD_PCR扩增和限制性酶切。这个简单、经济和可靠的改进方案也适用于许多其他的顽拗植物类群。 相似文献
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综述了国内外与植物抗虫性有关的次生物质的主要类型和植物次生物质对昆虫的寄主选择、取食和产卵等作用的研究进展,对次生物质在生态系统中的作用也作了介绍,并展望了植物次生物质的应用前景。 相似文献
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Mornkham T Wangsomnuk PP Wangsomnuk P Jogloy S Pattanothai A Fu YB 《Genetics and molecular research : GMR》2012,11(1):572-581
DNA extraction is an essential step for molecular analysis of an organism, but it is difficult to acquire a sufficient amount of pure DNA from plant tissue with high levels of phenolic compounds, carbohydrates, proteins, and secondary metabolites. Jerusalem artichoke (Helianthus tuberosus) has high levels of such substances. We compared five commonly used methods of extracting genomic DNA in tests made with leaves and seed of four Jerusalem artichoke genotypes: 1) modified method of Tai and Tanksley, 2) method of Doyle and Doyle, 3) method of Porebski, 4) modified method of ?torchová, and 5) Plant DNA Kit of Omega Bio-tek. The quality and quantity of extracted DNAs were assessed by photometric assay, electrophoresis on 1% agarose gel and a PCR-based technique. The modified method of Tai and Tanksley was found to be superior for both young leaves and seed. The quality of the extracted DNA was confirmed by sequence-related amplified polymorphism. This information will be useful for molecular analyses of Jerusalem artichoke and other related Helianthus species. 相似文献
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火龙果(Hylocereus undulatus)是近年发展起来的一种新兴热带水果, 其茎富含多糖、多酚及其它次生代谢物, 黏性极大, 很难从中提取高质量的DNA。特别是一年生以上的老茎, 目前尚未有较好的DNA提取方法。为了解决这一难题, 该研究对CTAB+Tris-HCl洗涤法进行了3种方式的改良。结果表明, “改进三”方法可不受取样时期和取样部位的限制, 从一年生以上火龙果茎中提取的DNA质量最好且不含黏性物质, 可用于酶切与分子标记等生化和分子生物学实验。该研究探索了一条较为理想的火龙果茎DNA提取方法, 值得推广应用。 相似文献
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火龙果(Hylocereus undulatus)是近年发展起来的一种新兴热带水果, 其茎富含多糖、多酚及其它次生代谢物, 黏性极大, 很难从中提取高质量的DNA。特别是一年生以上的老茎, 目前尚未有较好的DNA提取方法。为了解决这一难题, 该研究对CTAB+Tris-HCl洗涤法进行了3种方式的改良。结果表明, “改进三”方法可不受取样时期和取样部位的限制, 从一年生以上火龙果茎中提取的DNA质量最好且不含黏性物质, 可用于酶切与分子标记等生化和分子生物学实验。该研究探索了一条较为理想的火龙果茎DNA提取方法, 值得推广应用。 相似文献
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Mohammad S. Uddin Wenli Sun Xinhua He Jaime A. Teixeira da Silva Qi Cheng 《Biologia》2014,69(2):133-138
High quality genomic DNA is the first step in the development of DNA-based markers for fingerprinting and genetic diversity of crops, including mango (Mangifera indica L.), a woody perennial. Poor quality genomic DNA hinders the successful application of analytical DNA-based tools. Standard protocols for DNA extraction are not suitable for mango since the extracted genomic DNA often contains secondary metabolites that interfere with analytical applications. In this study, we employed an additional step to remove polysaccharides, polyphenols and secondary metabolites from genomic DNA extracted from young or mature leaf tissue; then a modified traditional cetyl trimethyl ammonium bromide (CTAB) method was applied. The use of 0.4 M glucose improved DNA quality and avoided contamination and browning by polyphenolics, relative to the traditional CTAB method. This is an easy and efficient method for genomic DNA extraction from both young and mature leaves of mango. The isolated DNA was free of polysaccharides, polyphenols, RNA and other major contaminants, as judged by its clear colour, its viscosity, A260/A280 ratio and suitability for PCR-based reactions. This modified protocol was also used to extract high quality genomic DNA from other woody perennials, including walnut, guava, lychee, pear, grape and sugarcane. 相似文献
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Abolfazl Barzegari Sepideh Zununi Vahed Sina Atashpaz Sajjad Khani Yadollah Omidi 《Molecular biology reports》2010,37(2):833-837
Various investigations have been so far performed for extraction of genomic DNA from plant tissues, in which the extracted
intact DNA can be exploited for a diverse range of biological studies. Extraction of high quality DNA from leathery plant
tissues (e.g., coniferous organs) appears to be a critical stage. Moreover, for some species such as Taxus trees, bioprocess engineering and biosynthesis of secondary metabolites (e.g., paclitaxel) is a crucial step due to the restrictions
associated with extinction of these species. However, extraction of intact genomic DNA from these plants still demands a rapid,
easy and efficient protocol. To pursue such aim, in the current work, we report on the development of a simple and highly
efficient method for the extraction of DNA from Taxus baccata. Based upon our protocol, interfering phenolic compounds were removed from extraction using polyvinylpyrrolidone and RNA
contamination was resolved using LiCl. By employing this method, high quality genomic DNA was successfully extracted from
leaves of T. baccata. The quality of extracted DNA was validated by various techniques such as RAPD marker, restriction digestions and pre-AFLP.
Upon our findings, we propose this simple method to be considered for extraction of DNA from leathery plant tissues. 相似文献
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Several DNA extraction methods have been reported for use with digesta or fecal samples, but problems are often encountered in terms of relatively low DNA yields and/or recovering DNA free of inhibitory substances. Here we report a modified method to extract PCR-quality microbial community DNA from these types of samples, which employs bead beating in the presence of high concentrations of sodium dodecyl sulfate (SDS), salt, and EDTA, and with subsequent DNA purification by QIAamp columns [referred to as repeated bead beating plus column (RBB + C) method]. The RBB + C method resulted in a 1.5- to 6-fold increase in DNA yield when compared to three other widely used methods. The community DNA prepared with the RBB + C method was also free of inhibitory substances and resulted in improved denaturing gradient gel electrophoresis (DGGE) profiles, which is indicative of a more complete lysis and representation of microbial diversity present in such samples. 相似文献
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In order to investigate the mutation characteristics and to further examine the genetic variation of mutant sunflower (Helianthus annuus) obtained in plants grown from seeds exposed to space conditions, only the mature tissues such as leaf and flower could be used for DNA extraction after mutation characteristics were confirmed. The rich contents of polysaccharides, tannins, secondary metabolites, and polyphenolics made it difficult to isolate high-quality DNA from mature leaves of sunflower according to previous reports. Based on the comparison of the differences in previously reported protocols, a modified method for the extraction of high-quality DNA was developed. Using this protocol, the DNA isolated from sunflower was high in quality and suitable for restriction digestion (EcoRI, HindII, BamHI), random amplified polymorphic DNA study and further molecular research. Therefore, the modified protocol was suitable for investigating the genetic variation of sunflower using mature leaf DNA. 相似文献
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DNA extraction from plants: The use of pectinase 总被引:3,自引:0,他引:3
Steven H. Rogstad Brian Keane Carolyn Howes Keiffer Fred Hebard Paul Sisco 《Plant Molecular Biology Reporter》2001,19(4):353-359
Several earlier protocols for extracting plant DNA or RNA do not work well for a variety of plants because contaminating substances
coprecipitate with the nucleic acids and, thus, are present even at the last DNA-hydration step. While DNA extraction protocols
have been published in which pectinase is employed to break down these contaminating substances, here we present an alternative
modified pectinase protocol that potentially uses fewer steps and avoids the use of ethylene glycol monoethyl ether and phenol.
DNA analyses results are described for 6 plant species demonstrating that the method works across distantly related plant
taxa. 相似文献
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Isolation of high-quality DNA from plants, especially plants from the Cerrado, is notoriously difficult because of polysaccharides and secondary compounds produced by plants from this biome. DNA isolation and its quality may be compromised by chemical defenses such as tannins and phenols. Quantitative plant defenses tend to have a cumulative effect, increasing in concentration during leaf development, reducing DNA quality extracted in mature compared to young leaves. We report the effect of leaf age on DNA extraction of Dimorphandra mollis. Our working hypothesis was that the young leaves have more DNA than old leaves of the same individual because chemical defenses accumulate in older leaves. Young and old leaves were sampled from eight mature trees as well as leaves from eight seedlings in the north region of Minas Gerais State. Genomic DNA extraction followed the standard CTAB procedure. DNA isolation was very successful from young leaves of 16 individuals of D. mollis. The extracted DNA exhibited high quality and the DNA quantity was also high, with an A(260)/A(280) ratio above 1.8, which is within the optimal sample range. In contrast, DNA isolation from old leaves was not successful. When the DNA was extracted from old leaves, the DNA was brownish, indicating contamination by phenolic compounds. These metabolites oxidize the DNA irreversibly, which hinders amplification of DNA by PCR by inhibiting the action of enzymes such as Taq polymerase. PCR performed with DNA from young leaves of D. mollis was successful and produced strong bands for RAPD markers. 相似文献
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The high quality of DNA template is one of the key factors to ensure the successful execution of polymerase chain reaction (PCR). Therefore, development of DNA extraction methods is very important. In this work, chitosan modified magnetic particles (MPs) were synthesized and employed for extraction of genomic DNA from genetically modified (GM) soybeans. The extraction protocol used aqueous buffers for DNA binding to and releasing from the surface of the MPs based on the pH inducing the charge switch of amino groups in chitosan modified MPs. The extracted DNA was pure enough (A(260)/A(280)=1.85) to be directly used as templates for PCR amplification. In addition, the PCR products were separated by capillary electrophoresis for screening of GM organisms. The developed DNA extraction method using chitosan modified MPs was capable of preparation of DNA templates, which were PCR inhibitor free and ready for downstream analysis. The whole process for DNA extraction and detection was preferable to conventional methods (phenol-chloroform extraction, PCR, and gel electrophoresis) due to its simplicity and rapidity as well as its avoiding the use of toxic reagents and PCR inhibitors. 相似文献