Resuspending and diluting primers

Your primer stocks come lyophilized (freeze-dried) from the company. It is best to resuspend them (in TE buffer or sterile H₂O) soon after they arrive. If you must store them for a period of time before resuspending, place them at -20 °C.

Resuspending primer stocks (100 µM)

  1. Quick spin your primer stock tubes.
  2. Label the lids of your primers using the Tough Spot labels provided in the PCR lab.

  3. Both on your primer tube and written on the primer data sheet that accompanies your primer order you will see the nmol quantity of your primer. To calculate out the amount of TE buffer you need to add to bring your primer to a 100 µM stock concentration, simply multiply the nmol quantity amount by 10.

    For example, if your primer’s nmol quantity is 36, you should add 360 µL of TE buffer to the tube. This will give you a final stock concentration of 100 µM.

  4. Add TE, gently mix the tube, and incubate 15 min at room temperature to resuspend.
  5. Quick spin your primer tubes.
  6. Store your primer stock tubes at -20 °C or make 10µM working aliquots by following the instructions below.

Making 10 µM working aliquots

  1. To make 10 µM working primer aliquots, you will need to label two 0.5 mL tubes per primer (one for PCR and one for sequencing).
  2. Add 180 µL of sterile H₂O to each 0.5 mL tube.

  3. Add 20 µL of 100 µM primer stock.

    Be sure you are adding the correct primer to your labeled tubes!

  4. Mix and spin.