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Double Digest using enzymes from diff companies


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#1 ChrissyFL

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Posted 04 May 2009 - 10:05 AM

I need to do a double digest with XhoI from Takara and EcoRI from NEB. What do I do about buffer compatibility? Will the EcoRI from NEB work with EcoRI buffer from Takara (will also work with XhoI)?

I'm crossing my fingers.... haha

Thanks!

#2 madrius1

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Posted 04 May 2009 - 10:42 AM

Look at the buffer composition and compatibility. You should find it easily on the NEB website.

#3 klinmed

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Posted 04 May 2009 - 01:37 PM

I need to do a double digest with XhoI from Takara and EcoRI from NEB. What do I do about buffer compatibility? Will the EcoRI from NEB work with EcoRI buffer from Takara (will also work with XhoI)?

I'm crossing my fingers.... haha

Thanks!


Takara recommend their buffer H for both enzymes. NEB suggests their unique "EcoR1 buffer"

Thus, do your double digest in either Buffer H or NEB "EcoR1 buffer".

This will work fine as the buffers have very similar salt concentrations.

Hope this helps

#4 T C

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Posted 05 May 2009 - 09:53 AM

Hey

If you are not satisfied with double digestions, just digest with the first enzyme, precipitate and digest with the other sequentially. Thats what I do for enzymes from different companies. Better delay at this stage rather than repeating the whole cycle. However, if buffers are compatible there is no reason why double digestion should not work.

Best,
TC

#5 ChrissyFL

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Posted 05 May 2009 - 10:57 AM

Thanks for the suggestions!




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