首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 78 毫秒
1.
原有的放线菌基因组DNA提取方法费时、费力、费用高 ,且对极端环境放线菌成功率较低。利用微波热振惊提取固体培养基表面放线菌菌落基因组DNA ,具有快速、简便、费用低廉等优点。所得的基因组DNA可作为PCR反应的模板进行 1 6SrRNA基因有效扩增。这为大量放线菌菌株的快速鉴别和系统分类创造了条件。  相似文献   

2.
一种简单、有效的适于PCR操作的放线菌DNA提取方法   总被引:15,自引:0,他引:15  
目的:利用改良酶法发展了一种从微量(几百微升)发酵液中快速安全的提取放线菌基因组DNA的方法。方法:利用溶菌酶破壁,蛋白酶K和SDS除蛋白,成功提取较高质量的放线菌基因组DNA,所得的DNA可作为PCR反应的模板进行16SrRNA等基因有效扩增。结果:能从海绵和土壤分离的放线菌中成功提取基因组DNA。结论:该方法操作简单、费用低廉、不使用酚、氯仿等有毒害作用有机试剂,非常适于长期从事放线菌操作的研究人员。为大量放线菌菌株的快速鉴别、高通量筛选和系统分类研究创造了条件。  相似文献   

3.
Chelex-100快速提取放线菌DNA作为PCR扩增模板   总被引:8,自引:1,他引:7  
旨在建立有效扩增16S rRNA基因序列的放线菌DNA快速提取的方法。采用Chelex-100法提取放线菌DNA,使用PCR扩增16S rRNA基因序列评价提取核酸的质量。结果显示,Chelex-100法能够在10 min之内从放线菌中快速提取DNA,所提取的DNA可以直接用于PCR扩增反应,PCR扩增产物电泳条带清晰,符合理论预期结果。因此,Chelex-100法提取放线菌DNA可以作为16S rRNA基因序列PCR扩增的模板,该方法具有经济、简便、快速的特点,适合于放线菌菌株大规模地筛选和分类鉴定。  相似文献   

4.
目的:建立从全血中提取基因组DNA的方法,并评价提取DNA的产量和质量.方法:分别采用传统酚-氯仿抽提法和试剂盒法制备基因组DNA,比较这两种方法提取DNA的产量、纯度、稳定性以及方法本身的优缺点、费用等.结果:试剂盒法简单快速,但其制备的DNA产率、纯度、稳定性低于酚-氯仿抽提法.结论:采用酚-氯仿抽提法可以提取较高质量的基因组DNA,适用于下游的分子生物学实验.  相似文献   

5.
许丽娟  马骁  王洋阳  王静  潘晴  刘梅 《生物磁学》2011,(20):3946-3950
目的:建立一种经济、快速且高质量提取人体外周凝血DNA的方法。方法:摸索最佳的匀浆条件,对外周凝血块进行匀浆,采用Ⅺ法对匀浆液进行基因组DNA的提取,通过凝胶电泳、单重PCR和多重PCR检测凝血基因组DNA的提取产量和质量。并分别与常规的凝血基因组DNA提取方法,即蛋白酶K消化法,以及提取抗凝血基因组DNA的Ⅺ法进行比较分析。结果:最佳的匀浆条件为:39000map,15秒。在此条件下提取的基因组DNA完整性好,纯度和产量与蛋白酶K消化法提取凝血DNA和KI法提取抗凝血DNA的结果相比,没有统计学差异。单重PCR和多重PCR也获得了理想的扩增结果。结论:与常规的外周凝血提取方法相比(蛋白酶K消化法),本方法节省了时间和成本,能快速、经济、有效地提取外周凝血基因组DNA,可用于后续的科研和临床诊断需要,解决了部分科研机构血液基因组DNA的样本来源问题。  相似文献   

6.
三种人全血基因组DNA提取方法的比较   总被引:1,自引:0,他引:1  
目的:比较改良酚一氯仿抽提法、盐析法、试剂盒法从人全血中提取基因组DNA的效果,以期建立一种快速、经济的提取高质量基因组DNA的方法。方法:分别用上述三种方法从人全血中提取基因组DNA,通过紫外分光光度计、琼脂糖凝胶电泳、聚合酶链式反应(PCR)、限制性内切酶酶切检测提取的基因组DNA的产量、纯度和质量。结果:改良酚一氯仿抽提法与试剂盒法提取的基因组DNA相比,DNA的产量有统计学差异,DNA的纯度无统计学差异,但试剂盒法提取的基因组DNA有较明显的降解现象:盐析法与改良酚.氯仿抽提法、试剂盒法相比,基因组DNA的产量和纯度都存在统计学差异,并且基因组DNA聚合酶链式反应(PCR)扩增的稳定性也明显劣于另外两种方法;三种方法提取基因组DNA均能进行限制性内切核酸消化。结论:改良酚一氯仿抽据取法是一种经济、快速、高效、稳定提取人全血基因组DNA的方法,适用于批量临床标本处理。  相似文献   

7.
氯化苄法提取染色体DNA   总被引:13,自引:0,他引:13  
本文介绍了一种简便、快速提取细菌和真菌的基因组DNA的方法一氯化苄法。使用氯化苄法抽提基因组DNA,不仅具有快迅、简便、耗资少的优点;而且得到的基因组DNA纯度高,质量好,可直接用于酶切、杂交和PCR扩增等用途。该法可用于多种细菌和真菌基因组DNA的提取。  相似文献   

8.
目的:探讨一种通厢的基因组DNA提取方法.方法:采用改良的膜法分别从动植物组织、外周血、细菌、细胞等标本提取基因组DNA,DNA样品经紫外吸收、琼脂糖凝胶电泳、PCR扩增和酶切进行榆测.结果:该方法提取的基因组DNA纯度较高,电泳条带清晰,DNA质量能满足下游分子生物学研究的需要.结论:该方法简便快速、适用范围广,是提取基因组DNA的一种有效方法.  相似文献   

9.
目的:建立一种以食用菌菌丝体和子实体为原材料的快速提取其基因组DNA的方法,从而提高基因组DNA的提取效率,为食用菌分子生物学提供便利.方法:分别以平菇黑平王的菌丝体和子实体为材料,快速提取其基因组DNA,并以此为模板进行ITS序列扩增.结果:采用方法提取的基因组DNA结构完整,无明显拖尾现象,浓度大约为10ng/μl,以此为模板能够获得预期的ITS条带.结论:该方法具有简便、快速、经济、无污染等优点,提取的基因组DNA适用于PCR反应等分子生物学研究,提高了提取效率,可作为高通量的提取方法.  相似文献   

10.
PCR鉴定时的微量DNA快速制备   总被引:7,自引:0,他引:7  
用基因组微量DNA快速提取方法,从胡萝卜转化再生株样品中快速提取了基因组DNA,并以此为模板进行胡萝卜转化再生株的PCR快速检测的结果表明,这是转基因时PCR检测的一种快速、简便、有效方法。  相似文献   

11.
Extraction of cellular DNA from human cells and tissues fixed in ethanol   总被引:4,自引:0,他引:4  
DNA can be extracted from ethanol-fixed lymphoid cells and tissues. The fixation procedure is simple and rapid, and the DNA extraction itself is the same as that normally used for fresh tissue or cells. DNA extracted from ethanol-fixed material is indistinguishable from DNA extracted from fresh samples based on its purity, its ability to be digested with restriction endonucleases, and its ability to specifically hybridize to DNA probes. The capability to extract DNA from ethanol-fixed cells and tissues eliminates the need for stringent handling and storage requirements of fresh or frozen specimens.  相似文献   

12.
We have developed a quick and low-cost genomic DNA extraction protocol from yeast cells for PCR-based applications. This method does not require any enzymes, hazardous chemicals, or extreme temperatures, and is especially powerful for simultaneous analysis of a large number of samples. DNA can be efficiently extracted from different yeast species (Kluyveromyces lactis, Hansenula polymorpha, Schizosaccharomyces pombe, Candida albicans, Pichia pastoris, and Saccharomyces cerevisiae). The protocol involves lysis of yeast colonies or cells from liquid culture in a lithium acetate (LiOAc)-SDS solution and subsequent precipitation of DNA with ethanol. Approximately 100 nanograms of total genomic DNA can be extracted from 1 × 10(7) cells. DNA extracted by this method is suitable for a variety of PCR-based applications (including colony PCR, real-time qPCR, and DNA sequencing) for amplification of DNA fragments of ≤ 3500 bp.  相似文献   

13.
Formalin removal from archival tissue by critical point drying   总被引:15,自引:0,他引:15  
Fang SG  Wan QH  Fujihara N 《BioTechniques》2002,33(3):604, 606, 608-604, 606, 610
The extraction of high-quality nucleic acid may be problematic in formalin-fixed tissues because of cross-linking between proteins and DNA. Old fixed tissue specimens do produce fragmented DNA (<1.2 kb), which is only used for PCR amplification. Here we show that high molecular weight DNA (>194 kb) can be successfully extracted from fixed tissue samples (16-70 years old) by gradual dehydration and critical point drying. The reliability of extracted DNA was measured by its ability to serve as a template for the amplification of mtDNA fragments (403 and 1198 bp) and an nDNA fragment (1844 bp). In addition, fingerprinting analysis was performed using DNA from fixed human tissue to ensure the ability of extracted DNA to hybridize with the DNA probe. DNA derived by this method can be subject to amplification, complete digestion by restriction endonuclease, and hybridization.  相似文献   

14.
DNA提取的应用与相关技术分析   总被引:22,自引:0,他引:22  
董明  宫月华  王兰  袁媛 《遗传》2003,25(2):205-207
为了分析影响提取DNA的有关因素,采用标准酚—氯仿抽提法和蛋白酶消化法提取DNA。结果表明,平均每mL全血可提取200 ~ 300μgDNA;新鲜组织及OCT包埋冰冻组织提取DNA,平均每0.2g组织可获得200~300μgDNA;直接将石蜡标本提取物制作模版,PCR扩增良好。从4种组织中均获得较为纯净的DNA;OCT对组织DNA无不良影响。 Abstract:To explore influence factors of DNA extraction,the protease and phenol-chloroform method was used to extract DNA in whole blood,fresh tissues,frozen tissues embedding with OCT and tissues embedding in paraffin.It results that 200~300μg DNA was extracted from 1ml whole blood or 0.2g fresh tissues or frozen tissue embedding with OCT.DNA extracted from paraffin specimen can be directly used in PCR amplification.Purity DNA can be extracted from four kinds of different tissues.OCT hasn't harmful effect on tissue DNA extraction.  相似文献   

15.
To investigate whether coelomic fluid secreted by earthworms can be a noninvasive source of DNA, we amplified and sequenced DNA extracted from the coelomic fluid and muscle tissue of eight worms. The sequences obtained using DNA extracted from both sources were identical. All cytochrome c oxidase subunit I (COI) mitochondrial DNA sequences, including those retrieved from GenBank, formed a monophyletic group of Metaphire sieboldi. The results indicate that we successfully extracted total DNA from coelomic fluid secreted by earthworm.  相似文献   

16.
扬子鳄鞣制皮革和鳞片的DNA提取方法   总被引:12,自引:0,他引:12  
史燕  吴孝兵  晏鹏  赵哲 《动物学报》2004,50(2):297-301
运用一种改进的提取方法 ,作者从鞣制皮革中成功地提取了总DNA ,同时还对尾尖皮、鳞片、盐腌生皮等皮质进行了DNA提取 ;用 12SrRNA基因扩增的通用引物、扬子鳄鉴别引物、微卫星引物及RAPD引物进行PCR扩增 ,并对部分扩增结果进行测序 ,以检验提取效果。结果证明 ,几种皮质标本都可提取出DNA ,其中尾尖皮和鳞片的提取效果较好 ,用四种引物都可扩增出明显亮带 ;盐腌生皮和鞣制皮提取结果也很好 ,并且用12SrRNA通用引物、扬子鳄鉴别引物扩增的亮带较明显 ,可进行扬子鳄皮质用品等的分子鉴定及部分序列的扩增和测序研究  相似文献   

17.
In the field of epidemiology, Genome-Wide Association Studies (GWAS) are commonly used to identify genetic predispositions of many human diseases. Large repositories housing biological specimens for clinical and genetic investigations have been established to store material and data for these studies. The logistics of specimen collection and sample storage can be onerous, and new strategies have to be explored. This study examines three different DNA sources (namely, degraded genomic DNA, amplified degraded genomic DNA and amplified extracted DNA from FTA card) for GWAS using the Illumina platform. No significant difference in call rate was detected between amplified degraded genomic DNA extracted from whole blood and amplified DNA retrieved from FTA™ cards. However, using unamplified–degraded genomic DNA reduced the call rate to a mean of 42.6% compared to amplified DNA extracted from FTA card (mean of 96.6%). This study establishes the utility of FTA™ cards as a viable storage matrix for cells from which DNA can be extracted to perform GWAS analysis.  相似文献   

18.
CELⅠ酶的粗提取及其活性检测   总被引:6,自引:0,他引:6  
韩锁义  杨玛丽  盖钧镒  喻德跃 《遗传》2006,28(9):1112-1116
CEL I酶是第一个从真核生物中提取的用于高效特异切割DNA双链碱基错配和DNA扭曲的内切酶, 因而也是TILLING技术中用到的一种关键酶。文章对CELI酶的粗提取及其活性检测进行了研究。错配切割实验表明, CELI酶在包含有G→A点突变的杂合双链中, 能有效地在错配位点进行切割, 并可以通过ABI377测序仪获得直观的检测结果, 从而可以用于TILLING分析。  相似文献   

19.
Purification of DNA from formaldehyde fixed and paraffin embedded human tissue   总被引:71,自引:0,他引:71  
The ability to isolate DNA from preserved human tissues would provide numerous experimental opportunities. In this report it is shown that DNA can be extracted from tissues prepared for routine histopathological examination (i.e., fixed with formaldehyde and embedded in paraffin). Although the extracted DNA is not intact, it is double stranded, cleavable with restriction endonucleases, and suitable for a variety of standard techniques used in molecular biology.  相似文献   

20.
目的:在生物浸出中,微生物群落结构分析有着重要意义,而群落分析的基础是提取纯度高、损失少的基因组DNA。为了解决这一问题,本实验通过比较两种较常用的DNA提取方法,煮沸裂解法和试剂盒法,寻找一种灵敏、快速、经济实用的制备浸矿细菌基因组DNA的方法。方法:分别用煮沸裂解法和试剂盒法提取6种浸矿菌的基因组DNA,从所提取的基因组DNA浓度、纯度、回收率和对PCR扩增反应的影响方面比较了两种方法的提取效果;用两种方法来处理不同浓度梯度的一种菌,通过实时定量PCR来比较两种方法的灵敏性。结果:相同处理量(108个)的革兰氏阳性菌(1株)、革兰氏阴性菌(4株)、古菌(1株)经两种方法提取的基因组DNA差异较大,煮沸裂解法所得的6组基因组DNA更纯,其OD260/OD280的值更接近1.8-2.0(纯DNA的OD260/OD280在1.8—2.0之间),前者所提DNA回收率最大可达后者的16.7倍;煮沸裂解法只需较少菌(102个)便能让实时定量PCR检测到所提DNA模板浓度,比试剂盒法灵敏。结论:两种方法提取的基因组DNA均可用于后续的PCR扩增,此外,前者提取的DNA浓度随细菌浓度增加而呈线性增大,而后者随茵浓度增大,所提DNA量增加有限,因此,在生物浸出中微生物基因组DNA的提取可直接采用简单快速的煮沸提取法,为实验节约成本和时间。  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号