LONG PCR AMPLIFICATION OF THE FVIII GENE INTRON 22 GENE INVERSION
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0.625 µl 10 mM dGTP
0.625 µl 10 mM deaza GTP
5 µl P/Q primer mix (each at 4 µM)
3 µl A/B primer mix (each at 2 µM)
1.875 µl 100% DMSO
Total amount (set pipette value)
9.13 µl (9.2 µl) 14.875 µl (14.9 µl)
Allow each reagent to thaw fully, mix and spin down before pipetting.
Ignore the spurious accuracy of these figures! - when multiplied in a master mix they will be accurate.
For the above scheme make a master mix of all ingredients (excluding each DNA sample) X the number of samples to be amplified, including controls. This minimises pipetting losses of reagents and should decrease any tube to tube variability between reactions. The above volumes assume the addition of approximately 1 µl of template DNA to each reaction. If different amounts of template are used then adjust the volume of water in master mix 1 accordingly.
