Restriction Enzyme Digest

Normal DNA digest with restriction enzymes is in a total volume of 50 ul (microliters.) You can do the digest in less vol. if, for example, you are only go to run it on a "minigel" (30ml agarose gel). In general digest 0.25 to 2 ug (micrograms) per digest.


"X" ul DNA. 0.25 to 10 ul

5ul 10x buffer. Use the correct buffer - see chart

"Y"ul dH2O. Final volume equals 50ul

1ul ENZYME (if you add multiple enzymes, final volume of enzymes should not exceed 1% of total volume).

1ul of restriction enzyme always is greater than 1 unit. It is usually 10 to 20 units. 1 unit is enough enzyme to digest 1 ug DNA.

-Incubate 37oC for 1 to 3 hours.

-Sometimes you need to add Mg++ to the reaction (in the form of MgCl) because you add a large volume of DNA. That is, if "X" is large > 20ul, you need to add Mg. A good rule of thumb is for every 20ul of DNA add 1ul of 10mM MgCl.

Writing for MAPK resource
MAP kinase resource community
MAP Kinase Wiki
MAP Kinase Conference 2014

Latest MAP Kinase news

The Biochemistry, Biology and Pathology of MAP Kinases II, September 11-12, 2014, Vilnius, Lithuania

It`s our pleasure to invite you to the international event "The Biochemistry, Biology and Pathology of MAP Kinases II". The conference is organized by Enterprise Lithuania together with Swiss...

Next-Gen Kinase Inhibitors Conference - Cambridge Healthtech Institute, June 17 - 19, 2013

This established and well-recognized kinase conference addresses reoccurring questions about selectivity, safety, resistance and novelty. After many years of research and therapeutic developments of...

Control of the cell cycle progression by the MAPK Hog1

Eukaryotic cells coordinate various intracellular activities in response to environmental stresses, activating an adaptive program to maximize the probability of survival and proliferation. Cells...

Ultrasensitive MAPK/Erk activation in absence of protein synthesis in Xenopus oocytes

The mitogen-activated protein kinase (MAPK) cascade in Xenopus oocytes exhibits an all-or-none, ultrasensitive response, which is believed to result from a positive feed-back loop. Here we describe a...

Batsheva de Rothschild Seminar on Biochemistry, Biology and Pathology of MAP Kinases

This abstract book contains most of the abstracts presented to the Batsheva de Rothschild Seminar on Biochemistry, Biology and Pathology of MAP Kinases. An Aharon Katzir-Katchalski Meeting and an...

Project Sponsors & Friends

Swiss Institute of Bioinformatics Journalism differently - journalism about mass media

Lithuanian Biotechnology Association Enterprise Lithuania

PAGEPress Publications - scholarly open access journals International Society for Biocuration

European Bioinformatics Institute

The Coins