Replicating RNA, useful, after reverse transcription, for analysis on microarrays, comprises conversion to cDNA then reverse transcription of this to form antisense sequences

摘要

Method for replicating RNA comprises conversion to cDNA then reverse transcription of this to form antisense sequences. Method for replicating RNA comprises: (a) generating a single-stranded DNA (A), by reverse transcription (RT) of an RNA, using a single-stranded primer (P1) of defined sequence, an RNA-dependent DNA polymerase (Pol1) and deoxynucleotide triphosphates (dNTP); (b) removing the RNA; (c) generating a double-stranded DNA (B) from (A) using a single-stranded primer (P2) that includes a box sequence, DNA polymerase (Pol2) and dNTP; (d) separating (B) into single strands; (e) converting these to double strands, using a single-stranded primer (P3) that includes the sequence of a promoter at its 5'-end and the same sequence as P1 at its 3'-end, Pol2 and dNTP; and (f) using an RNA polymerase (Pol3) and NTP to generate many RNA single strands having defined sequences at both ends. Independent claims are also included for the following: (1) Kit for the new process; and (2) Method for analysis of nucleic acid by using the new method to prepare RNA then analysis of this on a microarray.