Is Too Much Phosphatase Bad?

Ian A. York iayork at panix.com
Tue May 13 07:56:48 EST 1997


In article <01IISGAINNZM8WWXTN at MSSCC.MED.UTAH.EDU>,
Robert A. Schoenfeld <schoenfeld at MSSCC.MED.UTAH.EDU> wrote:
>
>Are there any deleterious effects when too much phosphatase is used - 
>i.e. ragged ends?

I don't know exactly what the damage is, but from experience
overphosphatasing will drasticly reduce ligation efficiency.  I used to
avoid phosphatasing at all costs, until (d'oh!) I went through the
calculations and realized that I was over-treating by a factor of 100 or
so because I was just using 1 ul; I had just assumed that, as with most
enzymes, a moderate excess was no big deal. When I started calculating the
correct amount, which meant diluting the phosphatase way down, the
problems disappeared. 

Ian

-- 
      Ian York   (iayork at panix.com)  <http://www.panix.com/~iayork/>
      "-but as he was a York, I am rather inclined to suppose him a
       very respectable Man." -Jane Austen, The History of England



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