| abstract |
A method of amplifying RNA, comprising the steps consisting in denaturing the RNA by heating a solution containing it, synthesizing a first strand of cDNA from an appropriate primer in the presence of a reverse transcriptase, denaturing l 'heteroduplex formed, synthesize the second strand of cDNA from a second primer in the presence of DNA polymerase, then subject the cDNA obtained to amplification cycles in sufficient number. Thanks to the discovery that certain reverse transcriptases which are not known to be thermostable are capable, when used in sufficient quantity, of withstanding a denaturation treatment of RNA by heat at a temperature greater than or equal to 60 ° C., and even to be able to operate, after this treatment, at high temperatures corresponding to sufficiently discriminating conditions, this process is characterized by the fact that said RNA solution contains, from the start, all the reagents and solvents necessary for said steps and that all steps are carried out in the same container without adding reagents or solvents. It is thus possible to operate, under satisfactory conditions, in a single manipulation step, which in particular avoids the risk of contamination.n n n Application in particular to the diagnosis of bacterial or viral infections, and genetic disorders, as well as to cell typing. |