Is BSA essential in our restriction digests?

[TRolandB]

We (London biohackspace) were planning to do a restriction digest on wednesday. It would be using Kpn1 on a PCR product for viewing on a gel. No further processing required. Having just looked at the info sheet for Kpn1 (from Sigma) I noticed that the enzyme buffer solution requires the addition of BSA.

We haven't used restriction enzymes before; is it worth leaving out the BSA or should we wait to get some before we proceed?

Will

[Mega]

I imagine it reduces star acivity and makes reaction quicker...

Here from NEB:

Bovine Serum Albumin (BSA) is supplied with some restriction enzymes to prevent adhesion of the enzyme to reaction tubes and pipette surfaces. BSA also stabilizes some proteins during incubation.

I read one instruction, where he said he always adds BSA because it doesn't harm the reaction, and it may improve it. 

If you don't use the DNA, it may be worth trying without? 

[Mega]

http://www.protocol-online.org/biology-forums/posts/14585.html

Hm, if you add a little more (say 7uL instead of 3)  that may make up the losses because of adhesion...  

[Nathan McCorkle]

hmm, found this:
How to prevent losses of protein by adsorption to glass and plastic ☆
C.H. Sueltera, b, M. DeLucaa, b
Analytical Biochemistry
Volume 135, Issue 1, November 1983, Pages 112–119

An improved procedure for reducing the loss of protein by adsorption
to glass or plastic surfaces is reported. For working with proteins at
the microgram level, the solvent is modified by adding glycerol (50%
final concentration) or Triton X-100 (0.2 mm final concentration).
Coating the plastic or glass surfaces with proteins such as bovine
serum albumin or other materials is not as effective; adding proteins
such as bovine serum albumin to the solvent is counterproductive.

http://www.sciencedirect.com/science/article/pii/0003269783907388

--
-Nathan

[Cathal Garvey]

Might be worth considering that BSA has some buffering activity, too,
but I imagine it's not a big deal in a dedicated reaction buffer.

[TRolandB]

Cheers for all the replies. We'll give it a go without BSA and see how it goes. May try the glycerol / triton method if it doesn't work!

[Mega]

Make sure to tell us how it worked ;) 

[william...@gmail.com]

Will do - has been postponed to saturday (We may have some BSA by then anyway, but we'll run regardless.)

On 21 February 2013 09:58, Mega <masters...@gmail.com> wrote:

Make sure to tell us how it worked ;) 

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[Andreas Sturm]

Great, thanks ;)