Restriction digest mapping

The extraordinary complexity of human genes and their products has encouraged the development of extremely high-resolution analytical methods.75 Capillary electrophoresis is competitive with slab gel methods, with resolution up to the order of about 1,000 base pairs for sequencing, sizing, and detection of mutation. Reversed phase HPLC is useful for restriction digest mapping and MALDI-MS up to about 1000 base pairs. 

A RESTRICTION MAP is used to identify and locate specific restriction sites on a given piece of DNA. The size of a fragment is determined by running the restriction digest on an agarose gel. Fragments separate by size—the smaller ones move farther toward the bottom of the gel. 

Electrophoretic analyses of restriction enzyme digests and construction of a restriction enzyme map (see Experiment 15)... 

Restriction-site mapping in DNA may be performed by end-labelling with polynucleotide kinase as above, and subsequent partial digestion with the restriction enzyme. An overlapping series of polynucleotides with a common labelled terminus is thus formed, and size analysis of these affords a restriction map. Polynucleotide kinase may also be used to assay the number of 5 -phosphorylated termini in DNA generated as the result of the action of restriction endonuclease. In the presence of... 

The chemical properties of the platinated DNA, termed M13-12A-Pt(-)-Stu I, were investigated by enzymatic, digestion and gel electrophoresis experiments. Platinum completely inhibits cleavage of the DNA by Stu I, as expected from the earlier restriction enzyme mapping studies. In addition, the cis-[Pt(NH3)2 d(pGpG) ] and cA-[Pt(NH3)2 d(pApG) ] intrastrand crosslinks were... 

FIGURE 11.33 Restricdon mapping of a DNA molecule as determined by an analysis of the electrophoretic pattern obtained for different restriction endonuclease digests. (Keep in mind that a dsDNA molecule has a unique nucleotide sequence and therefore a definite polarity thus, fragments from one end are distinctly different from fragments derived from the other end.)... 

The XE6 DNA was digested with several restriction enzymes and analysed via Southern blots using a probe derived fi-om the pgaW gene. A restriction map of the X.E6 clone revealed that the complete gene should be present on a 3.0 kb EcoRl fi agment (see Fig. 1). 

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