DNA labeling by nick translation reagents:
DNA for labeling (concentration c > 150 ng/µl)
modified nucleotides: Biotin-16-dUTP, Digoxigenin-11-dUTP, conc. 1nmol/µl (Boehringer Man....
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Typical 5'-kinase labeling reactions included the DNA to be labeled, [[gamma]]-32-P-rATP, T4 polynucleotide kinase, and buffer (3). After incubation at 37degC, reactions are heat inactivated by in....
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De-phosphorylation of DNA
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 (100m....
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DEPHOSPHORYLATION OF LINEARIZED DNA
ALKALINE PHOSPATASE(CIP) DIGESTION
Digestion of protruding 5'-ends
1. Precipitate digested DNA and resuspend in a 100ml of 1X CIP Buffer.
2. Add 1U CIP for ....
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DNA-Methyltransferase AssayThis protocol was written by Jean-Pierre Issa, based on Adams et al.* Kam-Wing Jair has made some useful shortcuts that work well if you are careful. Here is Kam-Wing Jair'....
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Bisulfite Modification of DNA
Materials and RegentsPrepare immediately before use10 mM hydroquinone (Sigma; #H 9003)
40.5% sodium bisulfite (Sigma; #S 8890, 8.1 g/20 ml ), dissolve 8.1 g sodium bi....
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DNA methylation is an epigenetic event that affects cell function by altering gene expression and refers to the covalent addition of a methyl group, catalyzed by DNA methyltransferase (DNMT), to t....
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1. Prepare reaction mixtures per 50ul Add Enzymes last, gently vortex mixture, quick spin liquid to bottom of tube:LabelBiotin-dUTP Digox-dUTP FITC-dUTP Texas Red dUTP 10X Biotin dNTP5 ul 0 ul 0 ul 0 ....
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solution I： 100 mM NaCl, 30 mM Tris.Cl （pH 7.8）, 10 mM EDTA
solution II：0.1 M NaCl，0.2 M蔗糖，10 mM EDTA
裂解液： 0.25 mM EDTA, 0.5 M Tris.Cl, （pH 9.2），2.5% SDS，10 mM ?-巯基....
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