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Construction of pool of interfering nucleic acids covering entire RNA target sequence and related compositions

  • US 20090099043A1
  • Filed: 11/21/2008
  • Published: 04/16/2009
  • Est. Priority Date: 07/23/2007
  • Status: Abandoned Application
First Claim
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1. A method for producing a pool of interfering nucleic acid (iNA) polynucleotides comprising the following steps:

  • a) partially digesting a DNA producing DNA constructs;

    b) ligating Loop-1 linkers to one end of the DNA constructs wherein the loop-1 linker contains a type II restriction enzyme recognition site and a type III restriction enzyme recognition site;

    c) cleaving the DNA constructs with a type III restriction/modification enzyme;

    d) ligating a Loop-2 linker onto an end of each of the DNA constructs, wherein said Loop-2 linker has a type II restriction enzyme cognition site producing DNA constructs that have a Loop-1 linker at one end and a Loop-2 linker at a second end of the DNA construct;

    e) digesting the DNA constructs with a type II restriction enzyme;

    f) ligating the DNA constructs into iNA expression vectors; and

    g) transfecting the transcription vectors into cells under conditions wherein iNAs are produced.

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