AgaroseGelElectrophoresisofDNA
1) Dissolve 1 g of agarose in 100 ml of 1X TAE or TBE buffer (gives a 1% gel). See note for making LMP agarose gel. 2) Cast the gel with the comb in place.3) Add 6X gel loading buffer to sample and load the samples into wells.4) Run the gel submerged in 1X TAE or TBE (30-60 min. at 100-150V)5) Stain the gel with ethidium bromide solution (10 µl per 100ml of buffer) for 10 - 15 min. 6) View on a long wave UV......阅读全文
PCR产物纯化方法
Purification of PCR Products in Preparation for CloningJoseph SambrookPeter Maccallum Cancer Institute and The University of Melbourne, AustraliaDavid
Restriction－Digest
Materials:Restriction enzymes of choice, such as BamH1 and EcoRIRestriction enzyme reaction buffer, such as MULTI-CORE (TM) (Promega)70 % Ethanol100 %
蛋白质电泳
蛋白质电泳（主要内容如下）One-Dimensional SDS-PAGETwo-Demensional SDS-PAGEProtein Electrophoresis in Agarose Gel Gel StainingRecipesOne-Dimensional SDS-PAGE·
DNA的酶学操作
DNA的酶学操作DNA Modifying Enzymes (Michael Blaber)Introduction to bacterial restriction/modification system. It provides very useful background knowledge
凝胶电泳(gel－electrophoresis)操作注意事项
1.缓冲系统：在没有离子存在时，电导率最小，DNA不迁移，或迁移极慢，在高离子强度的缓冲液中，电导很高并产热，可能导致DNA变性，因此应注意缓冲液的使用是否正确。长时间高压电泳时，常更新缓冲液或在两槽间进行缓冲液的循环是可取的。2.琼脂糖：不同厂家、不同批号的琼脂糖，其杂质含量不同，影响DNA的迁移
凝胶电泳（gel－electrophoresis）的注意事项
影响电泳分离的主要因素：待分离生物大分子的性质：待分离生物大分子所带的电荷、分子大小和性质都会对电泳有明显影响。一般来说，子带的电荷量越大、直径越小、形状越接近球形，则其电泳迁移速度越快。2. 缓冲液的性质：缓冲液的pH值会影响待分离生物大分子的解离程度，从而对其带电性质产生影响，溶液pH值距离
凝胶电泳（gel－electrophoresis）常见问题分析
琼脂糖凝胶电泳检测DNA时,跑出的带后面出现拖尾现象,什么原因造成的?参考见解： DNA带模糊：1、 DNA降解??避免核酸酶污染。2、 DNA上样量过多??减少凝胶中DNA上样量。3、 所用电泳条件不合适??电泳时电压不应超过20V/cm，温度＜30℃,巨大DNA链，温度应＜15℃，核查所用电泳缓
基于PCR技术的染色质沉淀分析1
INTRODUCTIONAfter chromatin immunoprecipitation (ChIP), different PCR-based approaches can be used to determine how much DNA is precipitated at a locu
ELECTROPHORESIS－OF－DNA－IN－POLYACRYLAMIDE－GELS
ELECTROPHORESIS OF DNA IN POLYACRYLAMIDE GELSGel SizesSmall: 165 x 130 mmMedium: 165 x 200 mmLarge: 165 x 260 mm5% Anal
Agarose－Gels－for－Single－Stranded－DNA
1. Prepare 50X TAE as:242 g Tris Base57.1 mL Glacial Acetic Acid100 mL 500 mM EDTA, pH 8.0600 mL ddH2OMix. Bring volume to 1 L. Autoclave.2. Mix the f
DNA－Purification－from－Agarose－Gels
1. Separate DNA fragments in an agarose gel cast with 0.5 mg/mL Ethidium bromide. Locate bands with a hand-held long-wave UV lamp.2. Slice the gel wit
基于PCR技术的染色质沉淀分析
INTRODUCTION After chromatin immunoprecipitation (ChIP), different PCR-based approaches can be used to determine how much DNA is precipitated at a loc
基因型分析
Randomly Amplified Polymorphic DNA (RAPD)Randomly Amplified Polymorphic DNA (RAPD) by (DNA KAFFE)RAPD analysis has been successfully used in mapping
PCR的下游应用
· Agarose Gel Electrophoresis of PCR Products (Robert H. Cruickshank)· Agarose Gel Electrophoresis of PCR Products (Immunology Resourc
Analysis－of－Proteins－using－Small－Format－2D－Gel－Electrophoresis
Preparation of protein samplesIntracellular virus proteinsThe following method has been developed principally for the analysis of intracellular protei
PCR的下游应用
・ Agarose Gel Electrophoresis of PCR Products(Robert H. Cruickshank)・ Agarose Gel Electrophoresis of PCR Products(Immunology Resource)
Competitive－RTPCR－Strategy－for－Quantitative－Evaluation－5
3. Characterization of the method precision and repeatability.a. Ensemble PCRs in the same conditions established before using quantities of target in
Gene－splicing－and－mutagenesis－by－PCRdriven－overlap－extension
实验概要 Extension of overlapping gene segments by PCR is a simple, versatile technique for site-directed mutagenesis and gene splicing.Initial
Preparation－of－Yeast－DNA－Embedded－in－Agarose－Plugs
Preparation of Yeast DNA Embedded in Agarose PlugsAnja van Brabant(adapted from Iadonato, S. P., and A. Gnirke. 1996. RARE-cleavage analysis of YACs.
重组DNA的分离、克隆与测序实验手册8
B. Midiprep double-stranded DNA isolationA midi-prep double-stranded DNA isolation has been developed to generate a sufficient amount of template DNA
Engineering－BioBrick－vectors－from－BioBrick－parts/Colony－PCR
MaterialsPCR SuperMix High FidelityVF2 primer (5''-TGCCACCTGACGTCTAAGAA-3'')VR primer (5''-ATTACCGCCTTTGAGTGAGC-3'')De
DNA克隆
DNA克隆（主要内容如下）· General Procedure· PCR Cloning· Subcloning· ET Cloning· Vector Preparation· Ligation Re
细胞支原体污染的PCR检测
支原体菌株来源：M.Arginini ATCC23838 精氨酸支原体M.FermentaneATCC19989 发酵支原体M.SalivariumATCC23064唾液支原体M.HominisATCC23114人型支原体M.OraleATCC23714口腔支原体M.HyorhinisATCC290
细胞支原体污染的PCR检测
支原体菌株来源： M.Arginini ATCC23838 精氨酸支原体 M.FermentaneATCC19989发酵支原体 M.SalivariumATCC23064唾液支原体 M.HominisATCC23114人型支原体 M.Ora
DNA电泳（agarose胶）操作方法
一、 试剂与材料：1、 琼脂糖2、 电泳缓冲液（1×TAE）3、 10mg/ml溴化乙锭4、 上样缓冲液5、 电泳仪和水平电泳漕6、 透射紫外灯7、 胶带纸 二、 操作方法 按1～2%的琼脂糖浓度（据DNA样品分子量不同而定）配胶100ml ↓ 置于微波炉内加热搅拌，至琼脂糖完全溶解
PCR实验指导与常见问题分析5
MgCl2 concentrationRelationship between MgCl2 and dNTP concentrationdNTP concentrations of about 200µM each are usually recommended for the Taq polyme
Phage－DNA
IntroductionThe phage lysate from the plate contains bacterial DNA and RNA, as well as phage DNA encased in the phage coat. The following procedure, d
基于PCR技术的染色质沉淀分析
INTRODUCTIONAfter chromatin immunoprecipitation (ChIP), different PCR-based approaches can be used to determine how much DNA is precipitated at a locu
TRFLP技术的优缺点
T-RFLP（末端限制性片段长度多态性）该技术在应用的过程中，肯定需要在实验条件上不断进行改进，而这种改进的好坏自然而然需要实验结果的验证。V. Grüntzig在2002年做了该工作的一部分，结果认为，在限制性酶切时，很有必要去除其中影响DNA的酶切过程，并且实验证明了具体的酶切时间。具有说服力的
TRFLP的优缺点
该技术在应用的过程中，肯定需要在实验条件上不断进行改进，而这种改进的好坏自然而然需要实验结果的验证。。V. Grüntzig在2002年做了该工作的一部分，结果认为，在限制性酶切时，很有必要去除其中影响DNA的酶切过程，并且实验证明了具体的酶切时间。具有说服力的结果如下： 1，T-RFLP出数据的