A research associate working for
Intragene Therapeutics prepares an investigative PCR reaction to
screen human tissue samples for specific mutations. He uses 55 ng
of genomic DNA from one tissue sample in a single PCR reaction, to
amplify a 1,246 bp fragment. The PCR protocol requires a final
concentration for two primers of 1.5 uM each, and a final
concentration of 180 uM for dNTPs, in a total PCR reaction volume
of 65 uL. During the PCR process, the fragment is amplified for 33
cycles. He quantifies the amount of PCR products to be 74
ng/uL.

4. What is the amount of amplification of the PCR product in his
reaction – that is, the fold increase in the amount of amplified
DNA segment? Answer in decimal notation.

5. If one of the primer stocks in his lab has a concentration of
50 uM, what volume (in microliters) does he add of this primer to
have the final concentration specified in the PCR protocol? Answer
to the nearest whole number.

6. The dNTP master mix available in the company lab has a
concentration of 12 mM total dNTP. What volume, in microliters, of
this master mix does he add to his PCR reaction to have the final
concentration specified in the PCR protocol? Answer to the nearest
whole number.

ANSWERS: (given answers, need help on how to get them)

4. 210,000,000

5. 2 uL

6. 1 uL