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Elution amount


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5 replies to this topic

#1 xyz74

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Posted 18 March 2010 - 06:55 PM

Hello,

I have a question about mega preps. I am doing a mega prep on my plasmid with the Qiagen kit. In the last step, it says to elute the DNA in a suitable volume of TE buffer. My question is what is that suitable volume? I usually elute in 500 ul and I am not sure if that is too high or too low.

Thanks.

#2 vladooo

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Posted 23 March 2010 - 08:42 AM

It does not matter. You can even dilute in 100 litres if you like. It only depends on what are you going to do with that DNA then. But 500 ul is optimal for most of the purposes.

#3 pito

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Posted 23 March 2010 - 08:47 AM

It all depens on the end concentration you want.

And in order to know that you need to do some testing.

I used 30Ál as elution total and I used ultra pure MQ and not the buffer

If you don't know it, then ask it! Better to ask and look foolish to some than not ask and stay stupid.


#4 vladooo

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Posted 23 March 2010 - 11:01 AM

Qiagen Mega gives you about 2.5 mg DNA. Volume of 30 ul is too low - it would be hard-to-pippete gel.
2.5 mg / 30 ul => 83 ug/ul
2.5 mg / 500 => 5 ug / ul
Choose whatever you like.

#5 pito

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Posted 24 March 2010 - 02:10 AM

ah yes I see my mistake I used a normal prep, not a mega prep.

sorry for the mistake.

You are 100% correct Vladooo

If you don't know it, then ask it! Better to ask and look foolish to some than not ask and stay stupid.


#6 xyz74

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Posted 24 March 2010 - 01:48 PM

Thank you for the replies!! I usually elute in 500 ul and get about 1.5ug/ul. A coworker said to try 3-5 mls of elution buffer. Hence the confusion.

Thanks.




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