Initially, a mixture containing 1,000 ng/ml total RNA, 1.0 ml l dNTP mix 10 mM, 1.0 ml of random hexamers 50 ng/ml and 8.0 ml of H2O DEPC diethyl pyrocarbonate was incubated for 5 minutes at 65° C. Then, after adding 9.0 ml of a solution containing 2.0 ml of RT 10x buffer, 4.0 ml of MgCl2 25 mM , 2.0 ml of DTT 0.1 M and 1.0 ml of RNase inhibitor, RNaseOUT, the mixture was incubated for 2 minutes at 25° C. After adding 1.0 ml of the enzyme SuperScript II, we proceeded to further incubation for 10, 50 and 15 minutes at 25° C, 42° C and 70° C, respectively.
Inicialmente una mezcla conteniendo 1.000 ng/ml de ARN total, 1,0 ml de dNTP mix 10 mM, 1,0 ml de random hexamers 50 ng/ml y 8,0 ml de H2O DEPC dietil pirocarbonato fue incubada durante 5 minutos a 65º C. A continuación, después de adición de 9,0 ml de una solución conteniendo, 2,0 ml de tampón RT 10x, 4,0 ml de MgCl2 25 mM, 2,0ml de DTT 0.1 M y 1,0 ml de inhibidor de RNase, RNaseOUT , la mezcla fue incubada por 2 minutos a 25º C. Después del agregado de 1,0 ml de la enzima SuperScript II, se procedió a nueva incubación de 10, 50 y 15 minutos a 25º C, 42º C y 70º C, respectivamente.
Inicialmente uma mistura contendo 1.000 ng/ml de RNA total, 1,0 ml de dNTP mix 10 mM, 1,0 ml de random hexamers 50 ng/ml e 8,0 ml de H2O DEPC dietil pirocarbonato foi incubada durante 5 minutos a 65° C. A seguir, após adição 9,0 ml de uma solução contendo, 2,0 ml de tampão RT 10x, 4,0 ml de MgCl2 25 mM, 2,0ml de DTT 0.1 M e 1,0 ml de inibidor de RNase, RNaseOUT, a mistura foi incubada por 2 minutos a 25° C. Após o acréscimo de 1,0 ml da enzima SuperScript II, procedeu-se nova incubação por 10, 50 e 15 minutos a 25° C, 42° C e 70° C, respectivamente.