Restriction endonuclease discovery

Arber and Smith Temin and Baltimore Khorana Discovered restriction endonucleases Discovery of reverse transcriptase in retroviruses Synthesis of first gene... 

One of the major obstacles to molecular analysis of genomic DNA is the immense size of the molecules involved. The discovery of a special group of bacterial enzymes, called restriction endonucleases (restriction enzymes), which cleave double-stranded DNA into smaller, more manageable fragments, has opened the way for DNA analysis. Because each enzyme cleaves DNA at a specific nucleotide sequence, restriction enzymes are used experimentally to obtain precisely defined DNA segments called restriction fragments. 

To incorporate fragments of foreign DNA into a plasmid vector, methods for cutting and rejoining of double-stranded DNA are required. Identification and manipulation of restriction endonucleases in the 1960s constituted a key discovery for cloning. Today, commonly used restriction enzymes are produced recombinantly and distributed widely. 

The discovery of restriction endonucleases was of crucial importance in sequencing DNA. These enzymes recognize a specific sequence of four to eight bases in double-stranded DNA and cleave the DNA at a precise point in this sequence. For example, the restriction endonuclease known as AIul cleaves the sequence AGCT between the G and C and the one known as Pstl cleaves the sequence CTGCAG between the A and the G. These restriction endonuclease enzymes provide a reproducible way to produce precisely defined fragments of an appropriate size for sequencing. 

The DNA content of the smallest genome such as that of the DNA virus SV40 is of the order of thousands of base pairs and the analysis of even such small genomes would be impossible if they could not be reproducibly fragmented into smaller pieces. The discovery of restriction endonucleases from several bacterial species provided the tool for the specific cleavage of DNA. Once cleaved, the fragments of different sizes could be separated electrophoretically, isolated, and analyzed. 

The development of recombinant DNA technology and the discovery and construction of vectors were instrumental in molecular cloning. Human DNA or other DNA could be readily obtained as fragments of desired lengths after treatment with restriction endonucleases. These fragments then could be inserted with the help of ligases into circular vector DNA, such as plasmid DNA, which has been linearized by a cut with the restriction enzyme at a unique site. 

One of the most important developments in the recent history of biochemistry is the discovery of restriction endonucleases (also called restriction enzymes). Restriction endonucleases are enzymes that catalyze the double-strand cleavage of DNA at specific base sequences. That is, restriction enzymes cut DNA strands at specific DNA sequences, generating fragments of specific sizes that can be separated and visualized on gel electrophoresis (Figure 25.6). 

The discovery of site-specific restriction endonucleases in 1970 and the development of efficient DNA sequencing methods in 1977 sparked a revolution in biology. On Oct 1,1990, the 15-year project to sequence the complete human genome was officially begun. ... 

There is a systematic way in which restriction enzymes are named (Smith and Nathans, 1973). The first three letters are in italics, and represent the genus and species names of the host bacterium. A nonitalicized letter may appear next, representing the particular strain from which the isolate was purified. The name will end with a roman numeral to indicate the order of discovery for multiple restriction enzymes arising from the same bacterium. For example, ficoRl was isolated from Escherichia coli RY13, while HindUl was isolated from Haemophilus influenzae strain d, and was the third enzyme isolated from this strain. (Note that the strain for HindUl can be found in older publications listed as Rd. The R is part of an R-M system, which denotes Restriction endonuclease versus Modification enzymes. The R and M strain designations are routinely dropped.)... 

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