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Abstract
<p class="first" id="d9116190e79">Gene splicing by overlap extension is a new approach
for recombining DNA molecules
at precise junctions irrespective of nucleotide sequences at the recombination site
and without the use of restriction endonucleases or ligase. Fragments from the genes
that are to be recombined are generated in separate polymerase chain reactions (PCRs).
The primers are designed so that the ends of the products contain complementary sequences.
When these PCR products are mixed, denatured, and reannealed, the strands having the
matching sequences at their 3' ends overlap and act as primers for each other. Extension
of this overlap by DNA polymerase produces a molecule in which the original sequences
are 'spliced' together. This technique is used to construct a gene encoding a mosaic
fusion protein comprised of parts of two different class-I major histocompatibility
genes. This simple and widely applicable approach has significant advantages over
standard recombinant DNA techniques.
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