Patent ID: 6794141
Filing Date: 2004-09-21
Classification: C12N,C12Q

Abstract:
A method of amplifying RNA sequences complementary to one or more than one target polynucleotide that is single stranded or made single stranded, comprisinga) forming double stranded cDNA templates containing sequences present in said target polynucleotide, wherein said sequences are operably linked to a promoter region, by i) annealing said one or more than one single stranded target polynucleotide with a first oligonucleotide comprising a primer operably linked to a promoter region to form a first complex, ii) synthesizing one or more than one first strand cDNA by reverse transcription of said first complex, iii) degrading first oligonucleotides not used in i) or ii) above with exonuclease activity, iv) annealing said one or more than one first strand cDNA, after denaturing the hybrid(s) of single stranded target polynucleotide and cDNA or degrading the single stranded target polynucleotide from said hybrid(s), with a plurality of second oligonucleotides comprising a random primer region to form a population of second complexes, and v) forming one or more than one double stranded cDNA templates from said population of second complexes with DNA polymerase activity; and b) transcribing said cDNA templates with an RNA polymerase capable of initiating transcription via said promoter region to produce amplified RNA (aRNA) containing sequences complementary to said one or more than one target polynucleotide.