Protocol Online logo
Top : Forum Archives: : Molecular Biology

Both the PCR primers in a vial. - Did anybody mix the diluted primers in single tube for use? (Mar/29/2006 )

Hi,
I want to know whether anybody stores both the diluted PCR primers in single tube? If no then why? I am curious to know the answer.
Hope we had good views.
Thanks in advance!
Niraj

-Niraj-

I always do that, I made to many manipulation errors, and the less pipeting, the less errors in my opinion.
And I have so many primers pair that it saves space in my PCR reagents box...

-Canalon-

Very good and efficient technique. Save them at the optimal concentrations for your reactions and its thaw and add...

This can be especially helpful for multiple primer pairs and HTS applications that require the use of multiwell plates. It saves a lot of time storing primers in a plate and being able aliquot ONCE from this plate into a plate of PCR mix.

-vasussci-

The problem with this is that you may want to use those primers for sequencing or in a different PCR amplification, and then you will not be able to, since you have mixed them. But if you are doing many reactions with the same set, it makes perfect sense.

-phage434-

QUOTE (phage434 @ Mar 30 2006, 11:59 AM)
The problem with this is that you may want to use those primers for sequencing or in a different PCR amplification, and then you will not be able to, since you have mixed them. But if you are doing many reactions with the same set, it makes perfect sense.

Thanks, how about diluting the initial stocks in the separate vials and reconstitute both the primers at 10 pmol in a single tube? If, we wish to sequence then we can use the main stock solution at any time.

-Niraj-

QUOTE (Niraj @ Mar 30 2006, 02:20 AM)
Thanks, how about diluting the initial stocks in the separate vials and reconstitute both the primers at 10 pmol in a single tube? If, we wish to sequence then we can use the main stock solution at any time.

Exactly my option. I keep 100X solutions of each primers. Every now and then I thaw them to prepare my mixed working solutions with all the necessary primers in one tube.

-Canalon-

I wonders why most people store the diluted primers in the separate tubes?

-Niraj-

I've always operated under a separate tubes principle. It increases the flexibility of the primers in case you want to add a different n-terminal or c-terminal tag with another primer, etc.

-Matt

-MisticMatt-