C - Chemistry – Metallurgy – 12 – P
Patent
C - Chemistry, Metallurgy
12
P
C12P 19/34 (2006.01) C12Q 1/68 (2006.01)
Patent
CA 2730761
The present invention is directed to a method for performing an RT-PCR for amplifying a target RNA comprising the steps of (i) cultivation of a population of adherent cells in a cell culture vessel (ii) lysis of said population of adherent cells which is supposed to contain said target RNA in said sample vessel with a lysis buffer comprising between 0.05 M and 1 M of a chaotropic agent (iii) adding reagents to said sample vessel which are necessary to perform a reverse transcription reaction such that the said chaotropic agent is present in a concentration of about 10 to 60 mM in said sample vessel, and reverse transcribing said target RNA and (iv) amplifying said first strand cDNA by means of subjecting said sample to multiple cycles of a thermocycling protocol.
La présente invention porte sur un procédé pour effectuer une RT-PCR pour l'amplification d'un ARN cible comprenant (i) la culture d'une population de cellules adhérentes dans un flacon de culture cellulaire, (ii) la lyse de ladite population de cellules adhérentes qui est supposée contenir ledit ARN cible dans ledit flacon d'échantillon avec un tampon de lyse renfermant entre 0,05 M et 1 M d'un agent chaotropique, (iii) l'ajout audit flacon d'échantillon de réactifs qui sont nécessaires pour effectuer une réaction de transcription inverse, de façon à ce que ledit agent chaotropique soit présent dans une concentration d'environ 10 à 60 mM dans ledit flacon d'échantillon, et la transcription inverse dudit ARN cible et (iv) l'amplification dudit ADNc de premier brin au moyen de l'opération consistant à soumettre ledit échantillon à de multiples cycles d'un protocole de cycle thermique.
Kubista Michael
Stroembom Linda
Zoric Neven
F. Hoffmann-La Roche Ag
Norton Rose Or S.e.n.c.r.l. S.r.l./llp
Tataa Biocenter Ab
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