共查询到18条相似文献,搜索用时 765 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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红豆杉属植物三种不同总DNA提取方法的分析比较 总被引:3,自引:0,他引:3
红豆杉属植物均为濒危物种,也是国家一级保护植物.以红豆杉属植物叶片为材料,利用三种不同的DNA提取方法提取总DNA,用分光光度计和琼脂糖凝胶电泳方法检测所得总DNA的得率和纯度,用PCR扩增的方法检测所得总DNA的质量,并对三种不同提取方法的结果进行了比较分析.结果表明:CTAB法提取的DNA纯度和得率均较高,可直接用... 相似文献
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富含多糖和次生物质的芒果子叶总RNA的提取 总被引:20,自引:0,他引:20
为了提取芒果子叶总RNA,去除次生物质和多糖等杂质的严重干扰,采用了正交试验快速优化提取方法。实验证明70%的丙酮能有效地去除大部分杂质,提取过程一天内可完成,RNA含量可达677μgg·FW,A260A280比值为195,RNA的纯度及质量适合后续反应。 相似文献
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三种土壤微生物总DNA提取方法的比较 总被引:3,自引:0,他引:3
本文对3种常用的土壤微生物总DNA提取方法Martin法、高盐改进法及试剂盒法进行了比较,并通过DNA得率、纯度及16S rDNA V3可变区的PCR扩增结合DGGE法(denaturing gradient gel electrophoresis),分别对3种方法进行评价.结果表明,3种方法提取的DNA均能满足土壤微生物多样性分析的要求.其中试剂盒方法操作简单,提取的DNA质量较高,但DNA得率较低且成本昂贵.Martin法和高盐改进法用时较长,DNA得率较高,纯度较低,但对后续PCR扩增和DGGE分析没有明显影响,且成本低廉. 相似文献
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何首乌总RNA提取方法的比较及改进 总被引:3,自引:0,他引:3
目的:探讨不同提取方法对提取何首乌总RNA的质量影响,寻找适于何首乌成熟叶组织RNA的提取方法。方法:以何首乌成熟叶组织为材料,采用SDS/酸酚法、常规CTAB法及改良的TRIzol试剂法分别进行实验,并对所提取RNA的质量进行验证。结果:采用3种方法都能提取出RNA,但质量差异较大。其中改良的TRIzol试剂法能有效抑制次生物质的影响,提取的RNA产量可达70-110μg/g,纯度高于其他2种方法,D260nm/D280nm值为1.85~1.97。结论:改良的TRIzol试剂法操作简便,提取的RNA完整性和纯度较高,可以满足下一步实验的要求。 相似文献
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为获得高质量的基因组DNA,分别采用传统酚-氯仿法、高盐法、试剂盒法和改进酚氯仿法提取香鱼肌肉基因组DNA。琼脂糖凝胶电泳检测结果表明,改进酚氯仿法提取的基因组DNA电泳条带整齐明亮且无降解。紫外分光度计测定DNA浓度和纯度,结果表明,改进酚氯仿法提取的鱼类基因组DNA浓度约为300μg/mL,A260/A280为1.80-1.86。用改进的酚氯仿法提取的DNA进行AFLP分析,扩增结果稳定,电泳条带清晰。综上所述,改进酚氯仿法能够获得高质量DNA,且可以用于进一步的分子生物学研究。 相似文献
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Eukaryotic non-coding DNA is functional: evidence from the differential scaling of cryptomonad genomes 总被引:5,自引:0,他引:5
Beaton MJ Cavalier-Smitht T 《Proceedings. Biological sciences / The Royal Society》1999,266(1433):2053-2059
Genic DNA functions are commonplace: coding for proteins and specifying non-messenger RNA structure. Yet most DNA in the biosphere is non-genic, existing in nuclei as non-coding or secondary DNA. Why so much secondary DNA exists and why its amount per genome varies over orders of magnitude (correlating positively with cell volume) are central biological problems. A novel perspective on secondary DNA function comes from natural eukaryote eukaryote chimaeras (cryptomonads and chlorarachneans) where two phylogenetically distinct nuclei have coevolved within one cell for hundreds of millions of years. By comparing cryptomonad species differing 13-fold in cell volume, we show that nuclear and nucleomorph genome sizes obey fundamentally different scaling laws. Following a more than 125-fold reduction in DNA content, nucleomorph genomes exhibit little variation in size. Furthermore, the present lack of significant amounts of nucleomorph secondary DNA confirms that selection can readily eliminate functionless nuclear DNA, refuting 'selfish' and 'junk' theories of secondary DNA. Cryptomonad nuclear DNA content varied 12-fold: as in other eukaryotes, larger cells have extra DNA, which is almost certainly secondary DNA positively selected for a volume-related function. The skeletal DNA theory explains why nuclear genome size increases with cell volume and, using new evidence on nucleomorph gene functions, why nucleomorph genomes do not. 相似文献
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Pérez Chaparro PJ McCulloch JA Cerdeira LT Al-Dilaimi A Canto de Sá LL de Oliveira R Tauch A de Carvalho Azevedo VA Cruz Schneider MP da Silva AL 《Journal of microbiological methods》2011,87(2):208-212
Multiple Displacement Amplification (MDA) of DNA using φ29 (phi29) DNA polymerase amplifies DNA several billion-fold, which has proved to be potentially very useful for evaluating genome information in a culture-independent manner. Whole genome sequencing using DNA from a single prokaryotic genome copy amplified by MDA has not yet been achieved due to the formation of chimeras and skewed amplification of genomic regions during the MDA step, which then precludes genome assembly. We have hereby addressed the issue by using 10 ng of genomic Vibrio cholerae DNA extracted within an agarose plug to ensure circularity as a starting point for MDA and then sequencing the amplified yield using the SOLiD platform. We successfully managed to assemble the entire genome of V. cholerae strain LMA3984-4 (environmental O1 strain isolated in urban Amazonia) using a hybrid de novo assembly strategy. Using our method, only 178 out of 16,713 (1%) of contigs were not able to be inserted into either chromosome scaffold, and out of these 178, only 3 appeared to be chimeras. The other contigs seem to be the result of template-independent non-specific amplification during MDA, yielding spurious reads. Extraction of genomic DNA within an agarose plug in order to ensure circularity of the extracted genome might be key to minimizing amplification bias by MDA for WGS. 相似文献
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The DNA contained by particles of densonucleosis viruses 1 and 2 were analyzed within the particle, and properties of DNA extracted from these particles were determined. The DNA appears to exist as a single-stranded molecule with limited secondary structure within particles, as assessed by spectral changes induced by formaldehyde, melting profiles, and circular dichroism studies. The single-stranded DNA had an apparent molecular weight of 1.9 X 10(6) to 2.2 X 10(6) as assessed by differences in the molecular weight of virus particles and top component and percentage of nucleic acid. DNA extracted from virus particles in low-salt buffers possessed properties typical of a single-stranded molecule. Double-stranded DNA could be extracted from virus particles under appropriate high salt and elevated temperature. The linear double-stranded DNA extracted from both viruses had a molecular weight of about 3.9 X 10(6) to 4.1 ZX 10(6) determined by neutral sedimentation and electron microscopy and an equivalent genome size determined by reassociation kinetics. About 87% of the DNA was homologous between the two viruses. 相似文献
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猪Ⅱ型圆环病毒豫A株的全基因组克隆与序列分析 总被引:13,自引:0,他引:13
参照国外发表的猪Ⅱ型圆环病毒(porcine circovirus type 2,PCV-2)全基因组序列,设计一对PCV-2特异性引物,用该室分离的PCV-2豫A株感染PK-15细胞,从中提取PCV-2复制型基因组DNA,并以之为模板进行PCR扩增.回收PCR产物,构建重组测序质粒T-PCV-2.测序结果表明,猪Ⅱ型圆环病毒豫A株的全基因组为1767bp,与GenBank收录的PCV-2国外分离株核苷酸的同源性可高达97%.序列分析表明,复制型豫A株的基因组包含10个读码框架,其中ORF1、ORF2是其两个最主要的读码框架,分别编码314、234个氨基酸.豫A株和PCV-1间的ORF1、ORF2的氨基酸序列同源性分别为85%、66%,与其它PCV-2毒株间的ORF1氨基酸同源性均在98%以上,而ORF2的氨基酸同源性为92%~97%. 相似文献
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Estimation of nuclear DNA content in plants using flow cytometry 总被引:5,自引:0,他引:5
Flow cytometry (FCM) using DNA-selective fluorochromes is now the prevailing method for the measurement of nuclear DNA content in plants. Ease of sample preparation and high sample throughput make it generally better suited than other methods such as Feulgen densitometry to estimate genome size, level of generative polyploidy, nuclear replication state and endopolyploidy (polysomaty). Here we present four protocols for sample preparation (suspensions of intact cell nuclei) and describe the analysis of nuclear DNA amounts using FCM. We consider the chemicals and equipment necessary, the measurement process, data analysis, and describe the most frequent problems encountered with plant material such as the interference of secondary metabolites. The purpose and requirement of internal and external standardization are discussed. The importance of using a correct terminology for DNA amounts and genome size is underlined, and its basic principles are explained. 相似文献