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Rolling circle synthesis of oligonucleotides and amplification of select randomized circular oligonucleotides

  • US 5,714,320 A
  • Filed: 02/23/1995
  • Issued: 02/03/1998
  • Est. Priority Date: 04/15/1993
  • Status: Expired due to Term
First Claim
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1. A method for synthesizing a selected oligonucleotide having well-defined ends comprising:

  • (a) annealing an effective amount of an oligonucleotide primer to a single-stranded circular template to yield a primed circular template, wherein the single-stranded circular template comprises (i) at least one copy of a nucleotide sequence complementary to the sequence of the selected oligonucleotide and (ii) at least one nucleotide effective to produce a cleavage site in the oligonucleotide multimer;

    (b) combining the primed circular template with an effective amount of at least two types of nucleotide triphosphates and an effective amount of a polymerase enzyme without the addition of auxiliary proteins to yield a single-stranded oligonucleotide multimer complementary to the circular oligonucleotide template, wherein the oligonucleotide multimer comprises multiple copies of the selected oligonucleotide; and

    (c) cleaving the oligonucleotide multimer at the cleavage site to produce the selected oligonucleotide having well-defined ends.

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