Making PCR plates

NOTE: These are the amounts that BIO uses, but I have found greater success with reducing the primer and using 7.5ul Forward Primer and 7.5ul Reverse Primer.


 * 1) Remove 2 cold plates from freezer.
 * 2) Remove the two primers that you will use in the reaction (a forward primer and a reverse primer) and PCR Master Mix tubes from freezer. Melt the liquid by holding the tubes in your hand.
 * 3) Add 12.5 ul forward primer, and 12.5 ul reverse primer to one PCR Master Mix tube, for a 96 well plate.
 * 4) Place an eight well strip onto a cold plate, and a skirted plate in the other cold plate. A plate that is “skirted” means that there is an edge around the plate.
 * 5) Pipette 131 ul of the mix (forward primer, reverse primer, and PCR Master Mix) into each well of the eight well strip.
 * 6) Pipette 10.5 ul of the mix into each well of a 96 well plate using a multi-channel pipette.
 * 7) Seal the plate with self-adhering plastic and store in the freezer until you are ready to use it.

PCR reaction mix 100 samples: Total volume: 1050ul
 * 10% trehalose 625ul
 * ddH2O 200ul
 * 10 X buffer 125ul
 * MgCl2. 62.5ul
 * dNTPs 6.25ul
 * Platinum Taq 6ul
 * Forward primer 12.5ul
 * Reverse primer 12.5ul

Each reaction is 12.5ul. If you are not working on an entire plate of samples, eight well strips can be used. Each individual tube then requires:
 * PCR Master Mix 10.25ul
 * Forward Primer 0.125ul
 * Reverse Primer 0.125ul
 * Template DNA 2ul

When you have your plate made, continue to the PCR Procedure.

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Updated April 27 2009