DNA实验技术

1)Prior to any ligation reactionyou should always run one gel in which purified insert and cut backbone are run side by sidepreferably beside a known amount of cut DNA.You can then use this to estimat ...

Hattoti's protocol adapted to cell culture :Hattori MTugores AVeloz LKarin MBrenner D (1990)A simplified method for the preparation of transcrip-tionally active liver nuclear extracts.DNA Cell Biol.Vo ...

1) The ligation mixture contains the following:vector (~100 ng)　　insert (equimolar or 2 or 3 X molar concentration of vector)　　water added to 18 µl2) Heat the mixture at 45 ℃ for 5 min. to melt any an ...

This method is after MarchukD.et al.1991Nucl.Acids Res.19(5)pp1154. You will need:10 x Taq buffer (Promega)Taq Polymerase (Promega)Phenol/chloroform mix100mM dTTPTE bufferAbsolute ethanol70% ethanol 1 ...

NH4Ac and EtOH precipitation of DNA Add NH4Ac (10M stock or solid) to the sample for a final concentration of 2.5M mix (spin at 4℃ transfer the supernatant to a new tube; optional spin for extra purif ...

PROTOCOL TO EXTRACT DNA FROM PARAFFIN BLOCKS1.Cut 10-20X10μm sections of formalin fixed paraffin samples into eppendorf tubes.2.Add 1 ml xylene mix incubate at 55℃ for 15mins. Release pressure spin ...

CTAB TECHNIQUE / Method / Schedule / Protocol FOR DNA ISOLATION / DNA EXTRACTION FROM PLANT LEAF / LEAVES SAMPLES(see also DNA RNA double isolation procedure if both DNA and RNA are needed)Reagents ne ...

DNA Fragment Purification from Agarose or AcrylamideFor 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 ...

Source: Contributed by Mohammad Reza Abbaszadegan et al. Abstract: Single strand conformational polymorphism (SSCP)is the most widely used PCR-based methods for point mutation detection. The abnormal ...

1、Oligo design（1）The 5' end (the homology arm) - choose 42 or more (we usually choose about 50) nts for the homology arms from the target DNA sequence simply according to where you want to insert the ...

一、In a microcentrifuge tube prepare a solution of linear DNA (25-50ng) in deionized water or TE buffer (10-35µl). 二、Add: （一）10X ligation buffer 5µl （二）50% PEG 4000 solution (for blunt ends only) 5µl （ ...

一.实验目的 1.掌握DNA指纹图谱技术的概念、原理和基本操作过程2.学习DNA的限制性酶切的基本技术3.掌握琼脂糖凝胶电泳的基本操作技术，学习利用琼脂糖凝胶电泳测定DNA片段的长度，并能对实验结果进行分析。二.实验原理1984 年英国莱斯特大学的遗传学家Jefferys及其合作者首次将分离的人源小卫星DNA 用作基因探针，同人体核DNA 的酶切片段杂交，获得了由多个位点上的等位基因组 成的长度不 ...

The preferred method of de-phosphorylation uses the buffer system described by Pharmacia for most of their restriction endonucleases. You will need:10 x OPA Buffer (100mM Tris.acetate pH 7.5 100mM mag ...

Prepare a ligation mix: ...

1.Crash Individual sand flies in 0.5 ml lysis buffer (50 mM NaCl 10 mM EDTA50 mM Tris-HCl pH 8).2.Incubate The sand fly homogenates with 100 ng/ml Rnase at 37 ℃ for 30 minutes.3.Incubate The cell lysa ...

1.Excise band of interest from a TAE gel; estimate volume by weighing the gel slice.2.Dissolve the gel slice in 3 volumes of NaI (from Geneclean kit)at 55E C for 5 min or until the gel dissolves.3.Add ...

Subcloning should be easy and fast and work every time. The following protocols minimize the number of manipulations required to prepare DNA fragments for ligations thereby both saving time and increa ...

Instructions Modified for Simpson Lab1.PCR: Perform a standard PCR reaction using taq polymerase. (Do not use tfl as the PCR product must have a 3' adenine overhang that only taq generates.) Include a ...

Linker Ligation (with T4 ) DNA LigaseIn a microcentrifuge tube prepare a solution of blunt ended dephosphorylated DNA (100-500ng) in TE buffer (5-7µl). Add 1-2µg of phosphorylated linkers in 5µl of TE ...

The overall sequence of events is:• Titer and plate out phage • Lift plaques onto filters and prepare them for screening • Make a probe • Hybridize the probe to the filters • ...