Calculate the amount of 10uM forward primer solution to add to a PCR reaction (25uL total reaction) to make a final concentration of 0.4uM forward primer in the reaction.

(a)  

Calculate the amount of water you need to add to make a final concentration of 70% ethanol solution by using 100mL of pure (100%) ethanol.

(a)  

You have ordered some PCR primers. They have arrived as dried (lyophilised) pellets. You have 30nmol of your forward primer. What volume of water should you resuspend the primer solution in to give a final stock concentration of 100uM?

(a)  

You have been given a cell line (CELL LINE A) at a concentration of 5 x 10⁶/mL. You want to extract DNA from only 1.6 x 10⁶ cells. Calculate what volume of cells you need to use for this experiment.

(a)  

You’ve now extracted the DNA (CELL LINE A) and want to determine the concentration. First you want to perform a 1 in 5 dilution using water. You need 150uL final volume to run on the spectrophotometer. How much of your DNA sample will you use?

(a)  

You run the diluted sample on the spectrophotometer and receive the following results (water blank has already been subtracted):

Sample: A₂₆₀= 0.315, A₂₈₀=0.180

Remembering that 1OD at A₂₆₀= 50ug/mL of double-stranded DNA, what is the concentration of your DNA sample (CELL LINE A)

(a)  

You are now going to set up a PCR. But first you need to dilute your sample to 40ng/uL in a total volume of 30uL for the PCR. What volume of your DNA sample (CELL LINE A @ 78.75ng/uL) do you need to use to make this dilution?

(a)  

You are now going to prepare your mastermix. In addition to your sample, you have an additional 2 DNA samples to test. You also plan to set up a negative control (containing water instead of DNA template). How many reactions will you prepare a mastermix for?

(a)  

Each PCR will consist of a final volume of 25uL. What volume of 10X PCR Buffer will you add to your mastermix to ensure there is a final concentration of 1X?

(a)  

What are the three steps involved in PCR in order?

(a)