Patent ID: 6794138
Filing Date: 2004-09-21
Classification: C12Q

Abstract:
A method for the amplification of nucleic acid, said method comprising:(a) contacting mRNA having a poly (A) tail with a primer comprising poly d(T) and a second different sequence; and, generatng a first cDNA stand from the mRNA strand by extending the primer by reverse transcriptase and the appropriate nucleotide under the appropriate conditions, which creates a first RNA:DNA duplex; (b) denaturing the first RNA:DNA duplex; (c) forming a double stranded DNA; (d) adding a first promoter to the DNA by hybridizing a first promoter primer to the single stranded DNA; (e) forming a double stranded DNA promoter region by adding the appropriate reagents; (f) producing multiple copies of RNA comprsing at least part of said second different sequence from the DNA strand comprising the promoter; (g) contacting said multiple copies of RNA with random primers; and, generating a first DNA strand from the RNA by extending the primer by reverse transcriptase which creates a second RNA:DNA duplex; (h) denaturing the second RNA:DNA duplex; and, adding a second promoter to the DNA by hybridizing a second promoter primer to the single stranded DNA; (i) forming a double stranded DNA promoter region by adding the appropriate reagents; and, (j) producing multiple copies of RNA from the DNA strand comprising the promoter.