Question

We received primers for use in our next PCR experiment. The primers arrived lyophilized (a dry powder) at 30 nanomoles per tube in a 2ml tube (the volume of the powder is negligible for this problem). The primers must be rehydrated (dissolved) in nuclease-free water. You have only 5 ml of nuclease-free water. We need 30 tubes for class containing 2 ul of primer in each tube at a concentration of 50 nM. Do you have enough for class? If no, how many tubes can you prepare? If yes, how do you prepare the tubes for class?

Answer 1

let`s first make 100 uM stock

1 M= 10^6 uM

1 mole=10^-9 nmole

molarity= 100 uM=10010^-6 M

number of moles= 30 nmole= 3010^-9 moles

molarity= number of moles/volume of the solution in liters

10010^-6 M= 3010^-9 moles/ volume of the solution in liters

volume of the solution in liters= 3010^-9/10010^-6

= 3 10^-4 L

1 L= 10^-6 uL

so 3 10^-4 L = 3 10^-410^6= 300 uL

so add 300 uL of nuclease free water to make 100 uM stock.

now let`s make working stock of 1 uM

stock concentrationM1= 100uM

concentration required M2= 1 uM

volume of stock required V1=?

final volume V2= 1 mL

M1V1= M2V2

V1=M2V2/M1

= (11)/100

= 0.01 mL

1 mL= 1000 uL

so 0.01 mL= 1 uL

add 1 uL from the stock solution to 999 uL of nuclease free water to make 1 uM working stock.

final concentration we need is 50 nM, and total volume required= 230= 60 uL

stock concentration M1= 1uM= 1000 nM ( 1 nM= 10^-3 uM)

concentration required M2= 5 nM

volume of stock required V1=?

final volume V2= 60 uL

M1V1= M2V2

V1= M2V2/M1

= 5 60/ 1000

=0.3 uL

take 0.3 uL from the working stock and dilute with 59.7 uL of nuclease free water.

yes, using the given volume of water and primer, we can make  30 tubes for class containing 2 ul of primer in each tube at a concentration of 50 nM.