Cleavage restriction enzymes

Nielsen P.E., Egholm M., Berg R. H., Buchardt O. Sequence specific inhibition of DNA restriction enzyme cleavage by PNA. Nucleic Acids Res. 1993 21 197-200. 

It is clear that both intact cell MALDI-TOF and PFGE have their limitations. PFGE analyses probes the chromosomal DNA of microorganisms for variations in the locations of specific restriction enzyme cleavage sites, while MALDI-TOF mass spectrometry of intact cells primarily examines abundant proteins such as ribosomal proteins35 and those associated with or near bacterial cell walls.58 In order for MALDI-TOF to detect a variation, a mutation must lead to noticeable changes in the expression of cell wall—associated... 

A third approach is to completely synthesize a gene for the protein under study. The sequence does not have to be exactly the natural one but can be made with restriction enzyme cleavage sites that permit easy excision and readdition of particular fragments. Synthetic fragments containing various mutations can then be grafted in at will. Genes of this type have been made for rhodopsin and related proteins (Chapter 23) and for numerous other proteins. 

The product in each case is 5 -[32P] labelled DNA plus unlabelled carrier RNA. The preparation is essentially salt-free and ready for either secondary restriction enzyme cleavage or strand separation. 

Restriction enzyme Cleavage reaction in DNA Organism used as source of enz5mie... 

The variations in DNA sequences between individuals as determined by differences in restriction enzyme cleavage patterns are known as restriction fragment length polymorphisms (RFLPs). RFLPs are manifestations of the unique molecular genetic profile, or fingerprint, or an individual s DNA. 

Nuclease cleavage (flap cleavage, 5 -nuclease cleavage, or restriction enzyme cleavage). Minisequencing or primer extension. 

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