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Dilution problem

Dilution

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#1 kaeterine

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Posted 12 July 2012 - 01:06 PM

Hi all,

I have a basic dilution problem... I need to send some samples for sequencing and i need to send in 500ng/ul of DNA in 9ul of sterile water. My stock DNA is 424.6ng/ul, now how do i dilute my stock to my desired concentration. please help me, pleaseeeeeee... I've try the calculations but I'm not sure if they are correct. Thank you

KA

#2 pito

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Posted 12 July 2012 - 01:31 PM

500ng/µl and if you need to send in 9µl, you will need 500ng X 9µl = 4500ng DNA for 9µl

You have 423ng/µl , so how much would you need?

4500ng in total, means 4500/423 = 10,63µl ... however you should be able to dilute this till you get 9µl .. which you cant, because you allready need >9µl to get 4500 ng...

So .. A problem...

But you could have seen this before even starting this calculation: you cant turn 423 ng/µl into 500ng/µl .....

You need to concentrate your samples more.... Its too diluted... or perhaps 423 ng/µl is still good enough?

eg: http://www.vivo.colo...manip/conc.html

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#3 gebirgsziege

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Posted 13 July 2012 - 12:23 AM

are you sure you need 500ng???? sounds quite a high concentration for me or are you trying to do a genome (in which case the 9µL would be a very tiny amount)?

Edited by gebirgsziege, 13 July 2012 - 12:24 AM.

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#4 fraffly

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Posted 25 July 2012 - 02:30 AM

Your concentration is too low, you can't dilute 424ng/ul to 500ng/ul, you'd have to concentrate it, for example by evaporation.
But in my experience, when sending samples for sequencing they always want a concentration much higher than the one they actually need for a successful sequencing, they like to play it save. I've regularly used concentrations 5 times lower than demanded and it always worked well.
I'd send it in the concetration you have, it'll work.
Plus, measurement of DNA is rarely precise anyway so you're in a good range.