I want to purify PCR product.
I used XBaI and KPnI enzymes.
Now I want to make reaction mixture.
From the universal buffer table it is showing that I have to use 1xM
What should I understand from this information? How to calculate?
Pranathi Meda
Usually suppliers will provide the buffer with higher concentration, for example 5X or 10 X. You need the final concentration on 1X in your reaction.
so if its 50 ul reaction and your buffer is 10 X use 5.0 ul to get 1X final concentration.
if its 5X then use 10 ul.
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Laurence Stuart Dawkins-Hall
- Agreed
- check supplier concentration and then dilute by this amount
- Most stand alone buffers containing mg and dNTPs which you then add primers and enzyme to at 5 x or 10x
- Most ready reaction mixes which include enzyme as well and therefore you just add water and primers to are 2x
- Check the side of your tube or for more info the web site which will have a data pdf explaining exactly how you do this
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Ashraf Abbas
check firstly the quality of DNA. concentrated and high quality purified DNA is prefereable for double digestion
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