plant protein precipitation

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ara_bidopsis22
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plant protein precipitation

 i have a crude extract from a plant.  it has low protein content and i need to concentrate it so i can do sds page. how do precipitate it by using ammonium sulfate? thanks for any help

kjosephs
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Are you sure you want to do

Are you sure you want to do this by Am. Sulf.  precip?  Really soluble proteins can be difficult to precip completely with Am. Sulf and its not so easy to remove the residual Am. Sulf. so that the gel runs properly. 
I have always done this by TCA precip. Its fast and works very well.  I posted a protocol in the protocols section of Scientist Solutions, the link is below.
www.scientistsolutions.com/science-protocols_displayprotocol_102a121b-76f1-4101-b38c-c8a92e850bb7_43b3934b-82ef-40db-8aef-0c2e734edad7.aspx

 
Kris
 
 

qiang wang
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I had hard time to resolve my

I had hard time to resolve my protein from TCA precipitation, if you want, may also try acetone, below is the link
 
http://www.scientistsolutions.com/science-protocols_displayprotocol_102a121b-76f1-4101-b38c-c8a92e850bb7_381187fb-bb05-4361-b306-551d4626895b.aspx

kjosephs
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TCA precipatation alone will

TCA precipatation alone will leave your samples in too much salt and the gel will not resolve, that is why you need to follow it with several acetone washes as the TCA precipitation protocol describes.  I have never done only acetone precipation, so I do not know how it compares to TCA.
 
 
 
 
 

Ivan Delgado
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In my experience the most

In my experience the most common method is TCA precipitation, which works very well. Another method that is good is ammonium sulphate, which as already mentioned need special care.
If you are interested in trying something different which works pretty well but may not be as efficient as TCA, you could try a methanol/chloroform precipitation. If you've ever purified genomic DNA using standard laboratory procedures (phenol/chloroform), you've done this (and you can visualize that the better you are at pipetting supernatants and getting close to solid interphases, the better you'll be at this protocol). For a protocol take a look at this page:
http://ivaan.com/protocols/145.html
Good luck

ara_bidopsis22
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ara_bidopsis22
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faint bands from aamonium

faint bands from aamonium sulfate. must try tca and acetone or phenol.
thanks!

By Nature
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