内切酶列表：EnzymesGeneratingBluntEnds
Asymmetric sequences are indicated by *.Single letter code:R = G or A; Y = C or T; W = A or T; M = A or C; K = G or T; S = C or G;H = A, C or T;V = A, C or G;B = C, G or T;D = A, G or T;N = G, A, T or C. RecognitionsequenceEnzymeAAT^ATTSspIAGC^GCTEco47IIIAG^CTAluIAGG^CCTEco147IAGT^ACTScaIATTT^AAAT SmiICACGTC(-3/-3)^*BtrICAC^GTGEco72ICACNN^NNGTGOliICAG^CTGPvuIICAYNN^NNRTGMslICCC^GGGS......阅读全文
内切酶列表：Enzymes－Generating－Blunt－Ends
Asymmetric sequences are indicated by *.Single letter code:R = G or A; Y = C or T; W = A or T; M = A or C; K = G or T; S = C or G;H = A, C or T;V = A,
内切酶列表：Enzymes－Generating－Blunt－Ends
Asymmetric sequences are indicated by *.Single letter code:R = G or A; Y = C or T; W = A or T; M = A or C; K = G or T; S = C or G;H = A, C or T;V = A,
内切酶列表：Enzymes－Generating－5protruding－Ends
Asymmetric sequences are indicated by *.Single letter code:R = G or A; Y = C or T; W = A or T; M = A or C; K = G or T; S = C or G;H = A, C or T;V = A,
内切酶列表：Enzymes－Generating－3protruding－Ends
Asymmetric sequences are indicated by *.Single letter code:R = G or A; Y = C or T; W = A or T; M = A or C; K = G or T; S = C or G;H = A, C or T;V = A,
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
PCR基本实验方法（五）
Cloning PCR ProductsT-A Cloning Strategy: Taq and other polymerases seem to have a terminal transferase activity which results in the non-templated ad
PCR基本实验方法（五）
Cloning PCR ProductsT-A Cloning Strategy: Taq and other polymerases seem to have a terminal transferase activity which results in the non-templated ad
Cloning－PCR－products－using－TA－vectors
Cloning PCR products using TA vectorsby Paul N. Hengen, Ph.D. *Methods and reagents is a unique monthly column that highlights current discussions in
EZBlunt载体克隆介绍
EZ- Blunt载体环形图谱和多克隆位点 EZ-Blunt载体克隆常见问题分析与处理
DEPHOSPHORYLATION－OF－LINEARIZED－DNA
DEPHOSPHORYLATION OF LINEARIZED DNAALKALINE PHOSPATASE(CIP) DIGESTIONDigestion of protruding 5'-ends1. Precipitate digested DNA and resuspend in a
Generating－stable－cell－lines－in－HEK293
Generating stable cell lines in HEK293Prior to transfection, it is recommended that you linearize your pcDNA gene construct. Linearizing will decrease
Generating－stable－cell－lines－in－HEK293
Generating stable cell lines in HEK293Prior to transfection, it is recommended that you linearize your pcDNA gene construct. Linearizing will decrease
CIP－Treatment
set up the following reaction:CIP RxnH2O7.8 ml10x cip rxn buffer2.0 mlDNA(e.g; 3 kb vector; 0.2 mg/ml; 2 mg total)10.0 ml(1 u/ml) CIP0.2 mltotal20.0 m
cDNA－Libraries
cDNA LibrariesIsolation of corresponding genetic informationInstead of synthesizing a desired gene, can we used the amino acid information to directly
RACE(rapidamplification－of－cDNA－ends)技术2
具体的实验步骤cDNA第一条链的合成：我们建议进行cDNA合成的对照反应，这样可以对样品的 cDNA的合成进行鉴定。加入各种试剂之后，在气浴中42度保温一个小时。注意： 在水浴或酒精浴中保温回减少反应体积，从而降低第一链的合成效率。将管放于冰上，以终止第一链的合成反应。直接进行第二链的合成。cDNA
RACE(rapidamplification－of－cDNA－ends)技术1
RACE技术的简介cDNA完整序列的获得对基因结构、蛋白质表达、基因功能的研究至关重要。完整的cDNA 序列可以通过文库的筛选和末端克隆技术获得。末端克隆技术是20世纪80年代发展起来的。RACE(rapid-amplification of cDNA ends)是通过PCR进行cDNA末端快速克隆
Selfcircularization－of－Linear－DNA
In a microcentrifuge tube prepare a solution of linear DNA (25-50ng) in deionized water or TE buffer (10-35µl).Add:10X ligation buffer 5µl,50% PEG 400
与II型核酸内切酶有关的几个概念
粘性末端：cohesive ends是指DNA分子在限制酶的作用之下形成的具有互补碱基的单链延伸末端结构，它们能够通过互补碱基间的配对而重新环化起来。平 末 端 ：Blunt end在识别序列对称处同时切开DNA分子两条链，产生的平齐末端结构。则不易于重新环化。同裂酶：isoschizomers 能
Cloning－of－small－RNAs－with－5’－phosphate－and－3’－OH－ends2
3’ Adaptor Ligation and PurificationHeat shock the RNA by putting at 90°C for 30 seconds. Snap cool on ice.Set up the 3’Adaptor ligation reaction in a
Cloning－of－small－RNAs－with－5’－phosphate－and－3’－OH－ends3
Load your precipitated PCR samples into 2 consecutive lanes so as not to overload the lanes. For each different sample, I would run a separate ladder
Cloning－of－small－RNAs－with－5’－phosphate－and－3’－OH－ends01
IntroductionThe following protocol describes a procedure for the purification and cloning of miRNAs and other small RNAs in the 20-30 nucleotide size
PCR
PCRPolymerase Chain Reaction1) Add the following to a microfuge tube:10 ul reaction buffer1 ul 15 uM forward primer1 ul 15 uM reverse primer1 ul templ
酶切反应注意事项
一、 建立一个标准的酶切反应目前大多数研究者遵循一条规则，即10个单位的内切酶可以切割1μg不同来源和纯度的DNA。通常，一个50μl的反应体系中，1μl的酶在1X NEBuffer终浓度及相应温度条件下反应1小时即可降解1μg已纯化好的DNA。如果加入更多的酶，则可相应缩短反应时间；如果减少酶的用
DNA标记
DNA标记（主要内容如下） DNA Labeling by Nick Translation Random Primed Labeling End-Labeling Purification of Labeled DNA Non-isotopic Labeling OthersDNA L
PCR的下游应用
· Agarose Gel Electrophoresis of PCR Products (Robert H. Cruickshank)· Agarose Gel Electrophoresis of PCR Products (Immunology Resourc
酶切反应的建议
酶切反应建议一、 建立一个标准的酶切反应目前大多数研究者遵循一条规则，即10个单位的内切酶可以切割1μg不同来源和纯度的DNA。通常，一个50μl的反应体系中，1μl的酶在1X NEBuffer终浓度及相应温度条件下反应1小时即可降解1μg已纯化好的DNA。如果加入更多的酶，则可相应缩短反应时间；如
PCR的下游应用
・ Agarose Gel Electrophoresis of PCR Products(Robert H. Cruickshank)・ Agarose Gel Electrophoresis of PCR Products(Immunology Resource)
重组DNA的分离、克隆与测序实验手册5
C. Random fragment end-repair, size selection, and phosphorylationSince both sonicated and nebulized DNA fragments usually contain single-stranded end
Observation－of－living－and－plasticembedded－chick－embryos
The development of chick embryos has been studied since Aristotle. It is one of the most intensely studied organisms. One reason for this is that ther
Polymerase－Chain－Reaction－(PCR)－cont.
Polymerase Chain Reaction (PCR) cont.Choice of Polymerases for PCROne of the important advances which allowed development of PCR was the availability