please correct - dilution question (Mar/14/2011 )

given: 10ug of 10ul DNA stock (or 100ng/ul DNA concentration)
need: dilute it so you have 1ug of 200ul final volume in the working stock (or 5ng/ul)

Say I use the entire 10ul.

I used C1v1=C2V2 and have: (x ng/ul)/(100ng/ul) *200ul = 10ul
x = 5 ng/ul
where 5ng/ul*200ul = 1000ng

So if I put 10ul of the 10ug DNA into 190ul water (200ul final volume) I'll have put in 1ug of the stock because 200ul*5ng/ul = 1ug

Is this correct?

just a question before even starting to calculate this: what is the given part?

I mean: 10ug of 10ul DNA stock (or 100ng/ul DNA concentration)
What does it mean?

10µg of 10µl ? What is that supposed to mean? Then you say 100 ng/µl.... ?

100ng per µl equals 1000 ng for 10µl... But 1000 ng is not the same as 10µg...

You have lost me allready at the given part...
Could you explain it?

You mean there is 10µg in your tube that contains 10µl ?

or?

If so: then you have 1µg for each µl or 1000ng/µl.

The way how you use that formula: I really dont get it...
You use the formula to calcutate 5ng/µl, but this was allready given ? Or not? (you mention it in the "need" part, meaning that the 5ng/µl is what you want in the end, so its "fixed" (read what I wrote at the bottom of this post)


In stead of "using" the formula (without thinking), try to think yourself (forget the formula) just think...

You are nog using the formula like you should.
(al least thats what I think, reading your post)



Anyway, about your formula:


I want an end volume of 200µl with an end concentration of 5µg/µl ==> how much ng is there then?
200*5 = 1000 ng

THus: 200*5ng = V1*C1 and what do you know of V1 and C1 ?
you know what the concentration is: 1000ng/µl ==> so how much V1 do you need to take ??????




Or do you really need to use the entire 10µl ??? if so , off course it would become a different excercice.

10µl would be V1 then and 1000ng/µl would be C1, C2 would be 5ng/µl (because this is the concentration you want) thus V2 would be ?

===> 10µl * 1000 ng/µl = 5ng/µl * V2 ==> V2 would be 2000µl, meaning you need to fill it untill you reach 2000µl .. But I have no idea what you are trying to do really...


Very simple: the formula you use is used to either calculate the amount of volume you need to ad to meet a certrain concentration with fixed starting volume/concentration (ex.2) or you use it to calculate a startvolume(=huw much you need to take from your "stock solution") you need to take to end up with a fixed volume of a certain concentration (ex1)

I'm not sure what it confusing. Maybe my wording is not right but I did not know there is a correct wording format. I always say 10ug of 10ul, or sometimes 10ul of 10ug. The given is saying that I have 10ul of DNA stock with amount of 10ug. This is also equivalent to 100ng/ul (10ug/10ul = 100ng/ul). I am just asking if I make a working stock where 10ul of the 100ng/ul is added into 200ul final volume (10ul DNA+190ul water), does this give 1ug final amount in the working stock? The working stock concentration is now 5ng/ul, so is it fine to do 5ng/ul*200ul = 1ug ?

claritylight on Mon Mar 14 20:22:46 2011 said:

I really dont understand it how you mention it.

Are you saying that you have a stock solution with a volume of 10µl that contains 10µg DNA ?

If so: then how do you end up with 100ng/µl ? 10 µg DNA in 10µl means 1µg for 1µl and 1µg is 1000ng ...

pito on Mon Mar 14 20:28:35 2011 said:

Think you are right, made a number error. So it's (10ug/10ul)*1000ng/1ug = 1000ng/ul

Then (10ul/200ul)*1000ng/ul = 50ngul

So I have 50ng/ul*200ul = 10000ng = 10ug

Is that ok

claritylight on Mon Mar 14 21:31:39 2011 said:

Its still not clear to me what you want.

As far as I understand it now you have a solution that containts 10µg DNA in a volume of 10µl (giving a 1000ng/µl stock solution) and you want to end with a working concentration of 5ng/µl in 200µl and thus you need to calculate how much of the stock solution you need ?Is this correct?

Or do you want to use the entire stock solution and dilute this untill you have a 5ng/µl solution?


I really still dont understand how you calculate that 10µg ... I dont get it... the 10µg is a given data... so why calculate it??

I really think you need to "forget" that C1*V1=C2*V2 formula for a moment and think!

What you need to do now is:

- write down what you know (the given data, and 10µg of 10µl means nothing...)
- write down what you want or need to solve. What is asked? Do you need to have an end concentration of 5ng/µl and need to use the 10µl (complete stock solution) untill its diluted till you reach that 5µg/µl or they want you to have a final volume of 2O0µl with a 5ng/µl concentration and calculate how much of the stock solution is needed for that?

Since we fixed my number mistake now, the given is 1ug/ul of stock DNA and I have 10ul so I have 10ug DNA amount right? I was just given the numbers of 10ug and 10ul

Just tell me if the below calculations are correct and I will figure out what I want on my own and we can all move on:

If I put 10ul of the 1ug/ul DNA into a tube, add 190ul water, this = 200ul total volume, and the DNA in this tube
now has concentration of (10ul/200ul)*1000ng/ul = 50ng/ul right?

What I want to know is how much amount of DNA is in the tube. I did 50ng/ul*200ul = 10000ng = 10ug, so there's 10ug of DNA in the tube. Is that right?

If you have 10 ug of DNA in a 200 ul volume, the concentration is 10ug/200ul = 0.05ug/ul = 50 ng/ul.

claritylight on Mon Mar 14 22:35:19 2011 said:

Ok, now I get it, see: telling you have 10µl of a 1µg/µl stock is not the same as just saying: 10µl of 10ng...



10µl of 1µg/µl in a volume of 190µl means you have in total 10µg in 200µl thus 10µg/200µl = 1/20µl = 1000ng/20µl = 100ng/2µl = 50ng/µl.


this is what homebrew allready did.


So yes, if you have 50ng/µl and you have 200µl , yeah, you have 50*200 ng DNA= 10000ng = 10µg...

But I dont understand why you do this second calculation!

You started by saying you would add 10µl of a 1µg/µl solution in that tube... So you knew from the start it was 10µg ..
(10µl*1ng/µl = 10ng .. and no matter how much water you add, even its 19999999liter, it will still stay 10ng of DNA in the end... Its the concentration that changes, not the amount of DNA itself)


You calculate B , using A to calculate A in the end ...

But still: I have the feeling you dont understand the formula or that you dont know what you are doing.