# some primer dilution calculations - (Mar/03/2008 )

hello all,

i have a very basic question, but am not able to solve it and need your help. I have a primer diluted at 1microgram/microlitre. now i need a 100micromolar solution or the current concentration of the solution in micromolar, can anyon help me with this conversion please. i really am stuck and not able to do anything about it.

regards

aditi

i have a very basic question, but am not able to solve it and need your help. I have a primer diluted at 1microgram/microlitre. now i need a 100micromolar solution or the current concentration of the solution in micromolar, can anyon help me with this conversion please. i really am stuck and not able to do anything about it.

regards

aditi

hi,

sorry but didnt get ur question. can u come again pls in detail.. sorry

hi

sorry, i will tell again.

i have a stock solution of primer with 1microgram/microlitre concn. but i need to know and use the molar concn. say 5micromolar/reaction, how do i convert it and how do i use the primer stock that i already have?

i have a very basic question, but am not able to solve it and need your help. I have a primer diluted at 1microgram/microlitre. now i need a 100micromolar solution or the current concentration of the solution in micromolar, can anyon help me with this conversion please. i really am stuck and not able to do anything about it.

regards

aditi

hi,

sorry but didnt get ur question. can u come again pls in detail.. sorry

sorry folks i think i am still not very clear.

i will go in details then.

it was a practice in our lab, to reconstitute the new oligos (that came lyophilised) in a stock concn of 1microgram/microlitre. we used to see the details of the number of micrograms of oligo we have recieved and then added the same microlitres of water, as an eg. if we recvd 300microgram of oligo, we would add 300microlitre of water to it. we never bothered to no the molar concn of the stock. but recently we are moving onto a different study where one of our main labs, insists on using micromolar concn, where they want our stock to be 100micromolar, and to then further dilute and use.25micromolar of it per reaction. we do not wish to discard our already reconstituted oligos, so i needed to know how do i go about converting the stock that we have in molar concn, so that we r able to use .25micromolar of oligo/reaction.

i hope i made it a bit more clear this time. i despearately seek your help and advice.

regards

aditi

you need to know the molecular weight of the primer, with that as M=moles/volume(L) ; (1mg/ml/MW)=mM now from this C1V1=C2V2 to dilute your primers at 100mM.

i will go in details then.

it was a practice in our lab, to reconstitute the new oligos (that came lyophilised) in a stock concn of 1microgram/microlitre. we used to see the details of the number of micrograms of oligo we have recieved and then added the same microlitres of water, as an eg. if we recvd 300microgram of oligo, we would add 300microlitre of water to it. we never bothered to no the molar concn of the stock. but recently we are moving onto a different study where one of our main labs, insists on using micromolar concn, where they want our stock to be 100micromolar, and to then further dilute and use.25micromolar of it per reaction. we do not wish to discard our already reconstituted oligos, so i needed to know how do i go about converting the stock that we have in molar concn, so that we r able to use .25micromolar of oligo/reaction.

i hope i made it a bit more clear this time. i despearately seek your help and advice.

regards

aditi

1 ug of 1kb DNA = 1.52 pmole, or 1pmole of 1 kb DNA - 0.66 ug. But this is for dsDNA, I think for ss primers you would have to divide by two. Lots of calculations! I would tell them screw it, use the old method for all of your old oligos, but then use the molar calculations for anything new you get.

Hi,

Here is a link where you can work "smart" than working "hard"...

http://www.graphpad.com/quickcalcs/ChemMenu.cfm

So to dilute your primer, u first have to calculate it in terms of micromolar(since ur desired concentration is in micromolar), or calculate ur desired concentration in micrograms and work out how much u would need... I cant do it for you now, since I dont have the formula weight of your primers. So if u use the link above u should be able to calculate/dilute solution at ease. Hope this helps... Good Luck....

The molecular weight of your primers will be approximately (length of oligo) * 330. Most of your oligos will likely be about 20 bp long, so the molecular weight would be around 6600 Da. So, 1 ug/ul would be 1/6600 Molar or around 150 uMolar. If you have 40 bp oligos, they will be around 75 uMolar. If you have oligos with large modifications (fluorescent, biotin) you will have to calculate the molecular weight. I would not agonize about this process -- a number within a factor of two will be close enough for all reasonable applications of your oligos.