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差减杂交法是目前广泛应用于差异性基因分离和鉴定的方法之一。简单地讲，差减杂交法是基于不同样品间特定核酸序列 ( 基因组 DNA 或 mRNA) 在数量 ( 等位基因数目或拷贝数或表达量 ) 上的差异，通过分子间的同源性杂交将存在数量差异性的序列分离出来。从研究对象和使用的基本材料来看，大致可以分为两类，即基因组差减杂交法和 mRNA 或 cDNA 差减杂交法。 1 ．原理 就是将不同来源的基因组D ...

原理： 此技术建立于PCR基础之上，使用一系列具有10个左右碱基的单链随机引物，对基因组的DNA全部进行PCR扩增，以检测多态性。由于整个基因组存在众多反向重复序列，因此须对每一随机引物单独进行PCR。 单一引物与反向重复序列结合。 使重复序列之间的区域得以扩增。 引物结合位点DNA序列的改变以及两扩增位点之间DNA碱基的缺失、插入或置换均可导致扩增片段数目和长度的差异，经聚丙烯酰胺或琼脂糖凝胶 ...

试验原理： 煮沸法是根据Holmex和Quigley(1985)的方法改进而成的。在基因工程中， DNA 分子的切割是由限制性内切酶来完成的。限制性内切酶能识别特定的 DNA 序列，在一定的条件下切断双链 DNA 。限制性内切酶的作用效率是受多方面因素影响的，如反应温度，缓冲体系，离子种类与浓度， DNA 的纯度和甲基化程度等。对 DNA 进行酶切时，首先要选择适合的缓冲液。对于单酶切，应选用该酶 ...

1.PCR技术 PolymeraseChainReaction聚合酶链反应 它是一种模拟天然DNA复制过程，在有DNA模板、DNA聚合酶（Taq酶）、RNA引物和四种dNTP的情况下，通过高温变性（90℃－95℃，1－2min）低温退火（25℃－37℃，1－2min）中温延伸（60℃－75℃，90sec-5min）这样反复循环的过程中，在体外扩增特异性DNA片段的分子生物学技术。 1）、PCR反应 ...

1. Depurination of DNA fragments: Wash gel in 0.25 M HCl 5' (small gels) 7' (large gels) 2. Denaturation: Wash gel in (0.5 M NaOH 1.5 M NaCl) for 20' (small) to 30' (large). 3. Neutralization: Wash in ...

Steve Hahn ...

1. Grind 20-50 frozen flies with a mortar and pestle in N2(l) and suspend in 2 ml Lysis Buffer. 2.Add Proteinase K to 100 mg/ml; incubate 1-2h @ 37℃ with occassional swirling. 3. Phenol extract GENTL ...

Protocol written by James Hermaundefined Methylation-specific PCR (MSP) is a simple rapid and inexpensive method to determine the methylation status of CpG islands. This approach allows the determination of ...

Materials: phenol:chloroform (1:1) chloroform 1.Add an equal volume of buffer-saturated phenol:chloroform (1:1) to the DNA solution. 2.Mix well. Most DNA solutions can be vortexed for 10 sec except f ...

CpG islands can be variable in length. Anyone know by what percentage range they can typically vary? I can't find much of any herlp on Google. Cheers Chris -Chris Harris- ---------------------------- ...

-Elaine Pinheiro and Aurora Burds Connor MIT Jan2006 This protocol works very well with Hybond-XL membrane from GE Healthcare (formerly Amersham). 1) Digest your genomic DNA using about 10 μg per r ...

Does somebody know how to increase DNA concentration from a too weakly concentrated DNA sample obtained with a Qiagen Extraction Kit (the elution buffer was ddH2O) ?? -BATY- ------------------------- ...

This procedure isolates DNA from agarose gels by filtration through a filter-paper column. The column is made in a 500 μl tube from a slurry of filter paper in TE buffer. Materials Whatman 3MM fil ...

1、用合适的酶切割DNA并电泳。 2、于紫外灯下从凝胶上切下所要的DNA带，装入含1×TBE缓冲液的透析代内，用夹子封好袋口，并检查有无漏孔。 3、将透析袋（纵长）与两极间的边线垂直放于电泳槽内电泳，4～5V/cm电泳至DNA贴紧袋壁。 4、取出透析袋，挤出凝胶块，吸出袋内液体放入1.5ml的离心管内，10000rpm离心5min。 5、吸出上清放入离心管内，加入等体积的饱和酚和等体积 ...

About 1/2 to 1 cm of tail should be cut from properly marked mice (using toe or ear clipping etc) about the age of 2-3 weeks. Place these tails into marked 1.5 ml Eppindorf tubes for processing; they ...

I recovered DNA from gel by Millipore's kitIt's simple.Cut the band from gel and centrifuge for 10 minutesbut the efficiency is disappointedonly 20% or so. Can anyone reccomend me other methods?Thanks ...

For fragments from 200 bp to 10 kb the agarose purification is ideal. For smaller fragments (20 bp to 400 bp) the acrylamide purification is preferred. Solutions Crush and Soak Solution 500 mM NH4 OAc ...

Hahn Lab，The Fred Hutchinson Cancer Research Center and Howard Hughes Medical Institute http://www.fhcrc.org/science/labs/hahn/methods/mol_bio_meth/BigDye Protocol.pdf ...

Preparation of C. elegans Genomic DNA Protocol by Andy Fires Lab Modified my Min-Ho Lee and Sudhir Nayak This protocol is based on the original Fire lab protocol for isolating genomic DNA from C. eleg ...

Important Notes Before Starting • New users are strongly recommended to read the QIAGEN handbook before starting the procedure. •Before using the kit for the first time dissolve the lyophi ...