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Enzyme Units RNaseA


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#1 Axolotl9250

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Posted 25 July 2012 - 03:54 AM

Hi,

I'm following a protocol which requires 20U of RNAse A to be added to a DNA extraction. I can't get my head around working out how much I need to add. The RNaseA I have is this one http://products.invi...roduct/12091021 and apparently it has an activity of not less than 70 U⁄mg material.
Searching the web all I've discovered is "1 enzyme unit (U) = 1 μmol min−1. 1 U corresponds to 16.67 nanokatals.[1]"

I'm thinking if I need 20 units absolute, I need to work that out as a concentration of my volume in microlitres, and then use C1xV1=C2xV2:

2ml of DNA+TE Buffer Solution
20U = 20/2000 = 0.01 Units per microlitre.

I'm assuming if 1ml more or less equals 1g then 1 microlitre more or less = 1 milligram.

70U/mg x V1 = 0.01U/ul x 2000

V1 = (0.01 / 70) x 2000 = 0.285ul

This may be completely wrong but I'm not sure how else to work this out.

Thanks,
Ben W.


#2 mdfenko

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Posted 25 July 2012 - 05:35 AM

this is not a c1v1=c2v2 problem. you are not looking for a final concentration. you are looking for an absolute amount.

you have a 20mg/ml solution of rnase and the specific activity is 70 units/mg (1400 units/ml). you want 20 units to put into your dna solution. 20/70=0.2857.

you, therefore, want approximately 0.29mg (20.3 units). 20mg/ml=1mg/50ul. 0.29x50=14.5ul.

you want to add ~14.5ul of your 20mg/ml rnase solution to your dna solution.

or you can calculate this way:

20units/1400units/ml=0.014286ml=~14.3ul

Edited by mdfenko, 25 July 2012 - 05:42 AM.

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#3 Axolotl9250

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Posted 26 July 2012 - 01:12 AM

Thanks! That's much easier than I was playing it up in my head I think.

#4 Xavi123456

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Posted 22 November 2016 - 11:27 AM

mdfenko

But, you put  14.3ul  of RNAse independently of volume of DNA solution?, for example I have 500uL of DNA. Should I put 14.3ul? I dont understand that calculation.
Thanks a Lot



#5 mdfenko

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Posted 23 November 2016 - 04:18 AM

mdfenko

But, you put  14.3ul  of RNAse independently of volume of DNA solution?, for example I have 500uL of DNA. Should I put 14.3ul? I dont understand that calculation.
Thanks a Lot

 

the calculation was not to determine final concentration, it was to add a specific mass of enzyme to the solution. this will slightly dilute the sample but will deliver the specific amount of enzyme


talent does what it can
genius does what it must
i do what i get paid to do




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