Do I have enough of each triphosphate?

[John C Calhoun]

Will a set of dATP, dCTP, dGTP, and dTTP at a 100 mM concentration with 10 μmoles of each triphosphate also contain at least 0.04 μl of each?

The conversions are foreign to me

[sunmoonstars]

Hi John,

I have a little trouble understanding your question. So I will take a guess, but please write back with follow-up questions if you have them.

A 100mM solution will contain 100mM of each of the four triphosphates. If you want to dispense 10um to use for a reaction, this is a 1:10,000 dilution, so you will mix 1 ul of the solution with 10 ml of your reaction mix.

Does that help?
Tonya

[John C Calhoun]

Hi, Thank you

I will be ordering a set of dATP, dCTP,dGTP, and dTTP from this website: https://www.promega.com/products/pcr/ro ... s-_dntps_/
If you scroll down past the individual triphosphates you will the sets. The one I want is the first set at a size of 10µmol each. I will be using 0.04µl of each for the reaction. Will ordering THAT sets be enough to use 0.004 of each?

[SciB]

John,

If you order the set of dNTPs at 100mM concentration and the kit contains 10 micromoles of each dNTP, then the volume has to be 100 microliters. You said you will use 0.04 uL for each reaction. For PCR we normally use 100 uM dNTPs so that would be a 1:1000 dilution of your 100 mM stock. You did not say what volume your reaction mix is but obviously you will have way more than enough dNTPs to use 0.04 uL per reaction.

You could take 1 uL of your 100 mM dNTP stock and make a 100-fold dilution using PCR grade water and then you could add 4 uL of this to each of your reactions. One microliter of your 100 mM stock gives you enough dNTPs to do 25 reactions so you definitely have enough to do a lot of experiments.

Let us know if you have more questions about the PCR mix or the amplification conditions.

Good luck!

Sybee