Reverse Transcription - (Jun/10/2005 )

The protocol:

10x RT buffer 2ul
25 mM MgCl2 4ul
0.1 M DTT 2ul
RNAaseOUT 1ul

Add 1ul (50 units) of SuperScript II RT to each tube, mix and incubate at 25°C for 10 min.


"25 mM MgCl2" is 25 mM the final concentration of MgCl2?


"Add 1ul (50 units) of SuperScript II RT to each tube" is 50 units the final concentration of SuperScript II RT?


Thanks!

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You need to give us the final volume of the reaction for us to give you the final concentration of the reagents.

Doh! since you use 2 ul of 10x buffer, your final volume is 20ul.

Your final concentration of Superscript is 2.5U/ul.

Your final concentration of MgCl2 is 25mM*4uL/20uL ... I'll let you do the math...I'm drinking beer.

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No, 25mM MgCl2 refers to the concentration of MgCl reagent solution that comes with your kit or working stock.

50 units refer to the total units in 1ul of SSII, thus also the number of units of SSII enzyme in one reaction

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