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Custom design for DNA snippers.

Custom design for DNA snippers

The powerful gene manipulations that underlie modern biotechnology rely on a set of chemical scissors called restriction enzymes. Normally produced by microorganisms, each of these enzymes snips DNA wherever it encounters a specific string of nucleotides, the DNA sub-units. A scientist can choose a restriction enzyme that cuts at one of about 100 different four- to six-nucleotide sequences. But there are many other segments where cuts might be desired.

Waclaw Szybalski of the University of Wisconsin at Madison now reports a method to customize a restriction enzyme to snip at any six- to 16-nucleotide sequence desired. It is as if a single adjustable wrench could replace, and provide more applications than, a toolbox full of fixed-size socket wrenches.

"It's a dream-come-true for enzymologists and molecular biologists who have always wanted a way to tailor their tools to particular tasks," says Szybalski. "Now we have a single, universal tool that can be easily adapted."

Whereas most restriction enzymes make their cuts at a site within the sequence of nucleotides that the enzyme recognizes, members of one small group of natural restriction enzymes have separate "recognition" and "cut" sites. For example, the enzyme FokI recognizes a stretch of five nucleotides, then cuts the DNA at a site a fixed distance away.

A hairpin-shaped piece of DNA, synthesized in the laboratory, serves as an adapter, allowing the scientists to snip a target piece of DNA at any specified site. One portion of the adapter contains the recognition site for FokI. The enzyme binds to this "portable" recognition site instead of requiring a site on the target.

The method takes advantage of the enzyme's requirement for double-stranded DNA at the cut site. Szybalski and his colleague Anna J. Podhajska include in the adapter a tail of single-stranded DNA, whose sequence specifies where the enzyme will cut. The enzyme FokI will make a cut only after the adapter tail forms links to a complementary sequence of nucleotides, found within the target DNA (provided in a single-stranded form). Once snipped, the DNA is released and reconverted to a double strand.

Some caution is expressed by IRA Schildkraut of New England Biolabs, the Beverly, Mass., company that sells FokI and that funded Szybalski's research. "As it stands, it's a clever thing to do, but it's not easy and it's relatively expensive," Schildkraut says. He sees its immediate value in special applications; for example, for cutting at sites with a relatively long specified sequence or for snipping DNA of large chromosomes into long pieces.
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Title Annotation:restriction enzymes
Author:Miller, Julie Ann
Publication:Science News
Date:Mar 15, 1986
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