红豆杉属植物三种不同总DNA提取方法的分析比较

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...     

一种优化的植物总DNA提取方法

根据自己提取植物基因组DNA的实际经验,改进了Clark利用CTAB提取植物基因组DNA的方法。在试验过程中发现,通过在研磨材料时加入液氮、用剪去端部的吸头转移含有DNA的溶液,并且将加入RNase A消化RNA的步骤改在最后进行等一系列改进,可以获得高质量的DNA。本文同时就主要提取步骤进行了分析。     

两种提取肠道微生物总DNA方法的比较

目的比较肠道微生物总DNA两种提取方法的优缺点。方法采用苯酚-氯仿抽提法和试剂盒法提取肠道微生物总DNA,比较其作为模板扩增细菌16S DNA的优缺点。结果两种方法提取的细菌DNA均能用于后续PCR扩增的模板。酚-氯仿抽提法经济可靠,但提取的DNA量及纯度较低试;剂盒法提取方便,操作简单,质量稳定,易标准化,但成本高。结论两种方法各具优缺点,应根据实验室条件和实验要求进行选择。     

鸡肠道菌群总DNA三种提取方法的比较

目的比较不同方法提取鸡肠道菌群总DNA的差异,为分子方法分析肠道菌群组成提供质量较高的DNA模板。方法采用反复冻融法、酶裂解法和试剂盒法（E.N.Z.A Stool DNA Kit）来提取鸡肠道菌群的总DNA,并根据DNA浓度及纯度、16S DNA扩增产物和ERIC-PCR产物所反映的片段多态性4个指标,对这3种方法提取的DNA质量进行比较。结果3种方法均能提取DNA,所得DNA都可以用于16S DNA的扩增,但后2种方法所得DNA的ERIC-PCR结果能反映出更高的菌群多样性。结论试剂盒法和酶裂解法所提取的DNA质量好,适合用于肠道菌群的分子生态研究。     

小鼠胚胎胃肠道细菌基因组DNA提取方法比较

目的:筛选能均衡地提取小鼠胚胎胃肠道微生物区系各种细菌总DNA的方法.方法:分别采用反复冻融法、CTAB -SDS法、柱式基因组DNA提取试剂盒法提取小鼠胚胎胃肠道细菌基因组DNA,对其进行琼脂糖凝胶电泳、紫外分光光度计测定、PCR扩增等质量检测.结果:CTAB -SDS方法提取的基因组DNA纯度较高,OD260/OD280平均值最高,为1.845,电泳条带清晰,能满足下游的PCR扩增等分子操作.结论:确定CTAB -SDS方法为提取小鼠胚胎胃肠道细菌基因组DNA的最佳方法,为研究不同种动物胚胎的肠道菌群的结构和多样性奠定了基础.     

三种提取蓝猪耳总DNA方法的比较

针对蓝猪耳（Torenia fournieri L.）叶片中多糖、色素等物质严重干扰总DNA提取质量的问题,以蓝猪耳叶片为试验材料,分别采用高盐低pH值法、SDS法、CTAB法提取总DNA,并从样品电泳图谱、纯度、得率等方面对三种方法进行评价。结果表明,CTAB法提取总DNA的产率较高、纯度最好,是蓝猪耳总DNA提取的最佳方法。     

活性污泥总DNA提取方法的研究

采用冻融＋玻璃珠、溶菌酶＋SDS和冻融＋玻璃珠＋溶菌酶＋SDS的方法提取活性污泥的微生物总DNA,并对以上三种方法进行比较,从中确定最佳的方法,以实现普通实验条件下成功提取符合PCR扩增要求的DNA。经紫外光度分析及电泳结果表明,冻融＋玻璃珠＋溶菌酶＋SDS方法所得的DNA的OD260/OD280为1．81,电泳结果显示,所提DNA片段分子量大于10kb,适于酶解和PCR扩增要求,为PCR技术应用于活性污泥的研究提供了一种简便、可靠的DNA提取方法。     

三种粪便总DNA提取方法的比较

目的比较不同粪便总DNA提取方法对肠道菌群多样性研究的影响。方法采用Bead beating法、化学裂解法和QIAamp DNA Stool Mini Kit提取同一份人粪便样品的总DNA,对比3种方法的DNA得率和16S rRNA基因V3区的变性梯度凝胶电泳（DGGE）图谱。结果Bead beating法的DNA得率约是其他2种方法的2倍;3种方法得到的DGGE图谱的Dice相似性为60％～70％,2条优势条带只出现在Bead beating法图谱中。在2～5min的Bead beating法击打时间里,DNA得率随击打时间的延长有一定的增加,但DGGE图谱无显著变化。结论不同的DNA提取方法会影响菌群的多样性分析。比较其他2种方法,Bead beating的裂解效率更高,能够检测到更多种类的细菌,更合适肠道菌群组成的分子研究。     

三种土壤微生物总DNA提取方法的比较

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.     

一种快速提取肠道微生物总DNA的方法

采集的兔肠道内容物及其粪便样品,通过分散浸泡、震荡洗涤、分级离心、滤器过滤、DNA提取试剂盒提取纯化,可以获得纯度很高的DNA样品。经0.8%琼脂糖凝胶电泳检测和紫外分光光度计测定,样品A260/A280的比值为1.72±0.02。分别以提取的DNA样品为模板,通过设计的细菌特异引物,对其16S rDNA基因进行PCR扩增,获得了1.6 kb大小特异性很好的预期条带。这为肠道微生物群落的分子生态学研究提供了一种简便、可靠的DNA提取方法。     

Evaluation of three methods for effective extraction of DNA from human hair

In this paper we evaluate three different methods for extracting DNA from human hair i.e. the Chelex method, the QIAamp DNA Mini Kit method and the ISOHAIR method. Analysis of DNA prepared from dyed hairs with the ISOHAIR method suggested that the DNA extracts contained PCR inhibitors. On the other hand, few inhibition was observed when DNA from dyed hairs were extracted using the Chelex method and the QIAamp DNA Mini Kit method. In conclusion, the Chelex method is recommended for PCR experiments in view of its simplicity and cost-effectiveness. To assess the reliability of the Chelex method for the extraction of genomic DNA from both natural and dyed hair samples, minisatellite variant repeat (MVR)-polymerase chain reaction (PCR) patterns of Chelex-extracted DNA were compared using hairs (three natural black hairs and three dyed hairs) with buccal swabs from six individuals. Complete agreement was observed between hair and swab samples in each individual, proving the utility of the Chelex method.     

Comparison of three methods of DNA extraction from cold-smoked salmon and impact of physical treatments

AIMS: To compare three bacterial DNA extraction procedures on cold-smoked salmon (CSS) and assess the impact on their efficiency of two physical treatments of the food matrix, ionizing irradiation and freezing. METHODS AND RESULTS: As molecular methods for bacterial detection have become an important analytical tool, we compared bacterial DNA extraction procedures on CSS. Working with frozen and irradiated CSS, we obtained negative responses from samples known to be highly contaminated. Thus, we decided to study the impact of these two physical treatments on bacterial DNA extraction procedures. The efficiency of bacterial DNA extraction directly from the fish matrix suspension was measured by an rpoB PCR-based reaction. The results demonstrated that the DNeasy tissue extraction kit (Qiagen, Courtaboeuf, France) was the most efficient and reproducible method. We also showed that freezing and ionizing irradiation have a negative impact on DNA extraction. This was found probably not to be due to inhibition as the PCR reaction remained negative after adding BSA to the PCR mix reaction. CONCLUSIONS: The extraction kit was the most efficient method. Physical treatments were shown to hamper bacterial DNA extraction. SIGNIFICANCE AND IMPACT OF THE STUDY: Attention must be paid to molecular bacterial detection on food products subject to freezing or to ionizing irradiation.     

Comparison of methods for total community DNA extraction and purification from compost

The differences on DNA yield and purity of three different DNA extraction protocols were compared with regard to the use for PCR and other molecular analyses. Total DNA was extracted from compost by the three protocols, and then was purified by spin-bind cartridges after being precipitated by PEG8000. The detection performed on a nucleic acid and protein analyzer showed that all three methods produced high DNA yields. The agarose gel electrophoresis showed that the fragments of crude and purified DNA had a length of about 23 kb. A eubacterial 16S rRNA gene-targeted primer pair was used for PCR amplification, and full length 16S rDNAs were amplified from all the purified DNA samples. After being digested by restriction endonucleases, the restriction map of amplified rDNA showed identical genetic diversity. The products of PCR using primer pair GC341F and 907R were also used for denaturing gradient gel electrophoresis analysis. The results indicated that high-quality DNA was extracted from compost by the three protocols, and each of the protocols is adapted to extract microbial genome DNA from compost expediently and cheaply.     

风沙土微生物总DNA不同提取和纯化方法比较

为筛选和建立风沙土中总DNA的提取和纯化方法,选取了5种直接提取法、1种间接提取法和2种纯化法分别对风沙土中总DNA进行了提取和纯化,并对其质量和产量进行了比较.结果表明：6种方法均可从风沙土中提取到大小为23 kb左右的总DNA,其中改进后的高盐提取法（用40%聚乙二醇8000和4 mol·L^-1 NaCl沉淀DNA）效果最好,纯化后总DNA的纯度最高,可进行16S rDNA的PCR扩增,且产量仅稍低于试剂盒提取法;电泳加柱回收纯化法的纯化效果较好,经该方法纯化后的总DNA大部分可进行PCR扩增,可满足后续分子操作对DNA纯度的要求.     

Comparison of three DNA extraction methods for Mycobacterium bovis, Mycobacterium tuberculosis and Mycobacterium avium subsp. avium

Aims:  To compare three methods for DNA extraction from Mycobacterium bovis , Mycobacterium tuberculosis and Mycobacterium avium subsp. avium .
Methods and Results:  The DNA was extracted from mycobacterial cultures using enzymatic extraction, combined bead beating and enzymatic extraction and cetyltrimethylammonium bromide (CTAB) extraction. The yield and quality of DNA were compared by spectrophotometry, agarose gel electrophoresis, restriction endonuclease analysis and PCR. The combined bead beating and enzymatic extraction method yielded more DNA. However, that method produced some sheared DNA, visible either by agarose gel electrophoresis or by restriction endonuclease analysis. All methods were appropriate for PCR amplification of a 123 bp fragment of IS 6110 in M. bovis and M. tuberculosis , and of a 1700 bp fragment of FR300 region in M. avium avium .
Conclusions:  Combined bead beating and enzymatic extraction method was the most efficient and easy method for extracting DNA from bacteria of the M. tuberculosis complex.
Significance and Impact of the Study:  The results reveal important differences among the DNA extraction methods for mycobacteria, which are relevant for the success of further downstream molecular analysis.