GE RPN1605 User Manual download pdf (Page 20)

5.3. Use of alternative reaction conditions

a. Use of more than one labelled [α–

P]dNTP.

Table 1 lists the results of a selection of standard reactions, using

a variety of input labels under optimum conditions. Figure 3 gives

more complete information on their use in Megaprime reactions.

Reactions were carried out at 37°C for 5 minutes.

b. Use of alternative specific activity [α–

P]dNTPs.

When using [α–

P]dNTPs of specific activity <3000 Ci/mmol the

incubation time should be extended to 1 hour at 37°C.

Figure 2. Time course of incorporation of [α–

P]dCTP (17 pmoles) in

a Megaprime reaction at 37°C. The DNA used was the standard DNA

supplied with the system.

% added label incorporated into DNA

0 10 20 30 40 50 60

Length of incubation in minutes

1 2 ... 15 16 17 18 19 20 21 22 23 24 25 ... 41 42

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GE RPN1605 User Manual Page 20