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1.grow up single colony in 1.5 ml LB/antibiotic ovn @37℃ .2.pour into tube spin for 30 sec .3.decant supernatant and resuspend pellet in 100 ml lysis solution .4.add 200 ml alkaline SDS vortex well .5 ...

This is a standard large scale prep.for plasmid DNA which gives a yield of 0.5-1.0mg.I have made some minor changes to the MHB protocol.Solutions Solution IIIIII from protocol D.1.Tris/EDTA pH 7.5 (op ...

Sample Preparation Cheek cells are obtained by rinsing the mouth with 25mls of any commercial mouth wash solution available for about 30sec the first thing in the morning. It is important not to brush ...

1、The BAC clone from glycerol culture sublibrary is innoculated into 3 ml of LB with 20µg/ml chloramphenicol and cultured at 37°C with shaking for 16hrs. The liquid bacterial culture is transferred in ...

A. Culture Growth: 1.For BAC isolation for shotgun library construction: Pick a smear of colonies and inoculate 3 ml TB medium containing the appropriate antibiotic. Grow the culture for 8 hours 37 de ...

1、A smear (rather than a single colony) of BAC colonies are picked and transferred into a 12X75 mm Falcon tube containing 3 ml of LB medium (10 g Bacto-Tryptone 5 g Bacto-yeast extract and 10 g NaCl i ...

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After an aliquot of the PCR mixture is analyzed on an agarose gel the remainder of the reaction is concentrated by ethanol precipitation resuspended in buffer and subjected to a simultaneous fill-in/k ...

Materials:3 M sodium acetate pH 5.2 or 5 M ammonium acetate DNA 100% ethanolMeasure the volume of the DNA sample. Adjust the salt concentration by adding 1/10 volume of sodium acetate pH 5.2 (final ...

This protocol gives very clean plasmid preps for restriction digests and cloning. However due to the alkaline lysis step the DNA is often nicked and may not give exceptional sequence data.SolutionsTEN ...

简介 优点：操作简单方便 缺陷：不能像EMSA或footprinting一样区分不同类型复合物 原理 双链 DNA 能够通过硝化纤维膜 (NC)，但是 DNA-protein 复合物不能通过NC filter. 通过定量分析留在滤膜上的复合物，可以判断二者的结合常数。 基本方法Lable DNA Mix DNA and protein to form complex Pass the mixtur ...

Pyrosequencing 在肿瘤基因甲基化研究的应用 甲基化研究对于基因的调控和肿瘤的发生有非常密切的关系。在一般正常的细胞中，基因调控区CpG岛处于非甲基化的状态。而当细胞发生癌变后，这些CG区域往往呈现甲基化状态。英国医学刊物《Lancet》报道奥地利因斯布鲁克医学院的专家们早就可以通过检测大便中DNA的甲基化变化，把健康人的大便与结肠癌患者的大便区分开。大便标本中SFRP2 甲基化的程度 ...

Molecularpourous membrane tubing is used for desalting protein and DNA isolation / purification. It comes in several diameters and pore sizes (for molecular weight conversions for nucleic acids see ge ...

Pellet 1.5 ml of an overnight culture at 12000 rpm in Eppendorf centrifuge at RT for 3 min. Resuspend bacterial pellet in 350 µl of STET buffer. Add 25 µl of freshly prepared solution of lysozyme (10 ...

Purification of DNA from agarose gels is an essential method involved in the sub-cloning of DNA fragments. The following method describes a variation of the method of Vogelstein and Gillespie 1979 (Pr ...

1.Obtain 65-100 µl of blood by retro-orbital bleed with a heparinized microcapillary tube. Expel blood immediately into a 1.5 ml microfuge tube containing 20 µl of 10 mM EDTA. Mix immediately to preve ...

This protocol is for Mini (up to 20 µg) preparations of high-copy plasmid DNA from cultures of E. coli.Important notes before startingNew users are strongly recommended to read Appendix A (page 21) at ...

作分子生物学实验，核酸纯化、酶切、克隆算是最基本的步骤了。因为要做定向克隆，通常要作双酶切。为了省时省事儿，我们常常想方设法把两个酶放在一起切。可是事情并非总那么称心如意――两个酶经常在一起闹别扭，不是反应温度不同啦，就是Buffer不同，有时候两个酶切位点连在一起，必须先切一个再切另一个，否则就切不动――因为有的酶除了识别位点之外还需要旁边留有几个碱基，如果正好被切掉了就出问题。于是有人就到处找 ...

1、将已经电泳确定的可回收的酶切产物在合适浓度的回收用琼脂糖凝胶进行电泳。最好换用新的电泳缓冲液10 × TAE（10 ×Tris-乙酸）。2、当溴酚蓝迁移至足够距离时（至少2 cm以上），在长波紫外灯下观察，用清洗过的刀片在目的片段前切下与目的片段同长，宽度适当（一般2 cm左右）的胶块。注意：①不要忘记在胶下垫一个新的塑料手套防止污染。②小心不要将回收胶切裂，同时注意与目的带相邻的切面要尽量平 ...

This is a modification of the procedure in Short Protocols (1-411-45)1. Prepare a 50 ml liquid lysate:A. mix 2xlO8 E. coli cells with 100 ul phage (from one picked plaque in 500 ul SM) and100 ul lOmM ...