Southern Blotting

The gel shift or electrophoretic mobility shift assay provides a simple and rapid method for detecting DNA -binding proteins. This method has been used widely in the study of sequence-specific DNA -bi ...

For use with GibcoBRL Random Primer DNA labeling system. Objective: To produce radioactively labeled DNA strands for the detection of target DNA or RNA sequences in various applications including So ...

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reagents: DNA for labeling (concentration c > 150 ng/µl) modified nucleotides: Biotin-16-dUTP Digoxigenin-11-dUTP conc. 1nmol/µl (Boehringer Mannheim) dNTPs (regular nucleotides): dATP ...

当双链DNA 分子的一条链上产生切口时,E.coli DNA 聚合酶Ⅰ就可将核苷酸连接到切口的3'羟基末端。同时该酶具有从5'→3'的核酸外切酶活性,能从切口的5'端除去核苷酸。由于在切去核苷酸的同时又在切口的3'端补上核苷酸,从而使切口沿着DNA 链移动,用放射性核苷酸代替原先无放射性的核苷酸，将放射性同位素掺入到合成新链中。最合适的切口平移片段一般为50-500个核苷酸。切口平移反应受几种因素的影响: (a) 产物的比活性取决于dNTP ...

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This protocol was designed to generate directionally end-labeled probes for DNaseI footprinting but it can be used for any application that requires end-labeled DNA probes. Solutions 10 mM dNTP Stocks Thaw100 mM stocks (NEB or Boehringer Mannheim) on ice and dilute 10-fold in Q. store small (10-20 ml) aliquotes at -80 degrees and thaw on ice just prior to use 10 X Klenow Buffer 0.5 M Tris 7.5 500 ml 1 M Tris 7.5 0.1 M MgCl2 100 ml 1 M MgCl2 10 mM DTT 100 ml 0.1 M DTT 0.5 mg/ml BSA 50 ml 10 mg/ml

Protocol for Annealing OligonucleotidesOligo Name:Lot Number:Total nmol:Volume of Annealing Buffer added:Oligo Name:Lot Number:Total nmol:Volume of Annealing Buffer added:Annealing Buffer: 10mM Tris, pH 7.5 - 8.0, 50mM NaCl, 1mM EDTA1xTE Buffer: 10mM Tris, pH 7.5 - 8.0,1mM EDTA.1.R ...

1. Prepare Whatman DE-52 according to manufacturers specifications. Equilibrate at store with 0.02% Sodium azide. 2. Plug a 1 mL (blue) pipet tip with Siliconized glass wool. Add 500 ml of 50:50 DE-52 ...

Wear gloves throughout and work in radiation area. Monitor area before and after use. Mix the following in an eppendorf tube: 1. 0.5 microgram oligonucleotide dissolved in H2O. 2. 3 microliters 10x kinase buffer. 3. 2 microliters 32 P ATP from ICN (>5000 ci/mmole). 4. H2O so that the final volume is 30 microliters. Add 25 units T4 polynucleotide kinase and incubate 60 min at 37 ℃. Purify labeled Oligonucleotide away from unincorporated ATP Currently, we use mini Quick Spin Oligo Columns (#1 8

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