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The Minion Lab College of Veterinary Medicine at Iowa State University http://mycoplasmas.vm.iastate.edu/lab_site/methods/DNA/SmaIvector.html ...

Has anyone used CTAB to remove polysaccharide during DNA extraction? I tried to use it under high NaCl but have had coprecipitation problem. My lysis buffer contains Tris EDTA NaCl and SDS. After solv ...

Table of Contents 10X TBE 40% Acrylamide Stock Alkaline lysis solution 10X TBE: 216 g Tris base 110 g boric acid 16.6 g EDTA Add water to 2 liters. 40% Acrylamide/Bisacrylamide (40% A&B): 38 ...

Purification of PCR fragments for cloning (adapted from Bruce A. Roe Department of Chemistry and Biochemistry The University of Oklahoma Norman Oklahoma 73019 broe@ou.edu) After an aliquot of the PCR ...

稀释限制性内切酶时，建议使用稀释缓冲液（A，B，C）。建议临用前稀释且稀释终浓度不要小于1000 units/ml。目录及说明书中标注了每一种内切酶相应的稀释兼容性。 注：以下稀释液不适用于耐热DNA聚合酶。欲了解稀释耐热DNA聚合酶的相关情况，请参见相应章节。 稀释缓冲液成份： 稀释液A： 50 mM KCl 10 mM Tris-HCl 0.1 mM EDTA 1 mM DTT 200 &m ...

Most recent version of protocol: 2/14/96 Dr. Paul J. Zambino Research Plant Molecular Pathologist U.S.D.A. Forest Service North Central Forest Experiment Station Forestry Sciences Laboratory 5985 Hwy. ...

Has anyone any experience of using a semi dry blotter for Southern blotting. At present I am tranfering overnight using a weight I think there is a blotter that is suitable for nucleic acids and can t ...

Quick question as I have read all sorts of views on this before: How long can bisufite-treated DNA be stored in -80℃ freezer before it degrades. Is there any reason why it should degrade quicker than ...

Reagents: Soln 1: 50 mM glucose/10 mM EDTA/25 mM Tris pH 8. Autoclave before use. Add 2 mg/ml Lysozyme just prior to use. Soln 2: 0.2 N NaOH/1% SDS. Keep solution at room temperature. Solution is norm ...

Acid Washing Beads Peter Novick Lab Department of Cell Biology Yale University School of Medicine http://info.med.yale.edu/cellbio/Novick/Second/Protocols/AcidBeads.pdf Materials: 1) 0.5 mm glass bead ...

The Preuss LabThe Division of Biological SciencesThe University of Chicago. http://preuss.bsd.uchicago.edu/protocols/enzyme.html ...

Preparation of electroporation cells 1. Prepare an overnight of NM522 in minimal medium. 2. Inoculate 1L LB with 10ml (1/100th vol) of the overnight and grow to A600 = 0.5- 1.0. 3. Pellet cells 5krpm ...

Growth and storage of Agrobacterium tumefaciens Strain GV3101: resistant to gentamycin and rifampicin so add 25-50 ug/ml Gentamycin 10 ug/ ml rifampicin on plates or in liquid media for selection. GV3 ...

重组DNA技术是现代分子生物技术发展中最重要的成就之一。即是基因工程(Gene Engineering)的核心技术。 重组DNA技术（Recombinant DNA Technique）是人类根据需要选择目的基因（DNA片段）在体外与基因运载体重组，转移至另一细胞或生物体内，以达到改良和创造新的物种和治疗人类疾病的目的。 这一技术的发展和应用，关键在于限制酶的发现和应用。 一、限制酶 限制性内 ...

(adapted from Bruce A. Roe Department of Chemistry and Biochemistry The University of Oklahoma Norman Oklahoma 73019 broe@ou.edu) Typically 2.5 - 3 volumes of an ethanol/acetate solution is added to t ...

DNA是通过异羟基洋地黄毒苷（digoxigenin，Dig）配基标记的脱氧尿嘧啶核苷三磷酸（dUTP）随机插入结合而被标记。dUTP通过间臂连结类固醇半抗原异羟基洋地黄毒苷酸基，形成Dig-dUTP，杂交反应后，杂交的靶DNA通过酶联免疫法与一个抗体复合物结合，接着在5-溴-4氯-3-吲哚磷酸盐（X-磷酸盐）和硝基四氮唑蓝（NBT）存在下，由酶催化反应，在杂交部位形成蓝紫色带或颗粒 ...

实验原理: Southern印迹是将DNA片断从电泳凝胶上直接转移至膜支持物（如硝酸纤维素膜、尼龙膜）上，使DNA片断固定的技术。先将DNA经限制性内切酶消化成一系列片段，进行琼脂糖凝胶电泳，各片段因分子量不同而彼此分开，然后经碱处理凝胶，使DNA的片段被变性、中和并通过毛细作用在高盐缓冲液中在原位将单链核酸转印到硝酸纤维膜上，烘干、固定。 试剂和器材 一、试剂 变性液：1.5mol/L NaCl ...

碱法提取的质粒DNA即使用RNA酶处理，仍会含有少量RNA。当有些试验需无RNA污染的DNA制品时，则需进行进一步纯化。一般常用Sepharose 2B或Sepharose 4B进行纯化，该方法具有快速，条件温和，重复性好，载体物质可以再利用等优点，因而已广泛用于质粒DNA纯化。 1、将Sepharose 2B经含0.1% SDS的TE（pH8.0）平衡后上柱。 2、将至多1ml的DNA溶液铺 ...

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 ...

TaKaRa的内切酶和NEB的内切酶哪个更好一些？ 参考见解： TaKaRa的内切酶、NEB的内切酶两个公司的酶的品质都非常好。NEB公司的酶的活性很高，切出两条小带可能是因为出现星活性，可以试试把酶量减半。NEB的酶很多都是克隆的，所以纯度比较高。 应用磁性微球提取人全血基因组DNA，将提取出的DNA直接用于限制性酶切反应，应用Taq1酶，但发现酶切后产生的是smier片断，即切碎的状态。不知 ...