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Method of inserting unique DNA sequences into DNA vectors

DC
  • US 4,528,266 A
  • Filed: 09/30/1982
  • Issued: 07/09/1985
  • Est. Priority Date: 10/01/1979
  • Status: Expired due to Term
First Claim
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1. A method of inserting a unique oligonucleotide sequence into a non-essential region of an autonomously replicating DNA element, comprising,isolating double stranded circular molecules of the DNA, each having the same nucleotide sequence,cleaving the circular DNA at random with respect to nucleotide sequence, producing a population of linear, double stranded DNA molecules comprising circular permutations of the same nucleotide sequence,joining a unique oligonucleotide sequence to the ends of the linear DNA molecules, said unique oligonucleotide sequence not otherwise existing in said DNA element, thenrejoining the ends to form circular double stranded DNA molecules having the oligonucleotide of unique sequence inserted at random with respect to the nucleotide sequence of each circular DNA molecule,transferring the circular DNA having said unique insert sequence to a host organism under conditions permitting replication of the DNA, andselecting for progeny of the circular DNA having a unique insert sequence, said progeny bearing said insert in a non-essential region of the DNA.

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