Electrophoresis endonuclease

Endonuclease-catalyzed hydrolysis of DNA at the internucleosomal linker regions into multimers of 180 base pairs which are visualized by electrophoresis as a ladder of nuclear DNA fragments. Access of the endonuclease to DNA is facilitated by depletion of polyamines, and the activity of the enzyme is mcrea.sed by and decreased by ADP-tibosylation. Thus, agents that increase intracellular Ca " or inhibit l>oly(ADP-ribose) polymerase can induce apoptosi.s. ... 

Treatment widi restriction endonuclease Agave 2 fragments, one 7 kb in size and one 3 kb in size, as judged by gel electrophoresis. 

Use of pulsed-field gel electrophoresis (PFGE) to separate large DNA fragments Permits isolation of large DNA fragments obtained by use of restriction endonucleases (rare cutters) that result in very limited cutting of DNA. 

Another popular technique for endophyte identification is RFLP (restriction fragment length polymorphism).In this technique, nuclear or mitochondrial DNA is extracted from the endophyte and digested with restriction endonucleases. The digestion products are separated by gel electrophoresis, and a unique pattern emerges for the fungus, similar to the patterns seen in the RAPD-PCR technique. 

Once a DNA molecule has been cleaved into fragments, a particular fragment of known size can be enriched by agarose or acrylamide gel electrophoresis or by HPLC (pp. 92, 90). For a typical mammalian genome, however, cleavage by a restriction endonuclease usually yields too many different DNA fragments to permit isolation of a particular fragment by electrophoresis or... 

Southern blotting is a technique that can be used to detect specific genes present in DNA. The DNA is cleaved using a restriction endonuclease, the pieces are separated by gel electrophoresis and then transferred to a nitrocellulose membrane for analysis. The fragment of interest is detected using a probe. 

Restriction patterns produced by agarose electrophoresis of DNA fragments after restriction endonuclease action. Courtesy of Bio-Rad Laboratories, Richmond, CA. 

Restriction endonucleases catalyze the hydrolysis of specific phosphodi-ester bonds in double-stranded DNA. These enzymes are often used to linearize a circular plasmid for hybrid DNA construction. They have also found use in the analysis of DNA and the construction of restriction maps. In this experiment, students will incubate various restriction enzymes with plasmid or viral DNA and analyze the product DNA fragments by agarose gel electrophoresis. 

You just isolated a novel recombinant clone and purified the desired insert (a 10,000 bp linear duplex DNA) from the vector. Now you wish to map the recognition sequences for restriction endonucleases A and B. You cleave the DNA with these enzymes and fractionate the digestion products according to size by agarose gel electrophoresis. Comparison of the pattern of DNA fragments with marker DNAs of known sizes yields the following results ... 

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