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Team:Cambridge/protocols/Norgen
From 2011.igem.org
Norgen Proteospin Inclusion Body Prep.
A proprietary kit for isolating the non-soluble fraction of a cell's internal protein (a soluble internal fraction is also obtained but usually discarded).
Theory
Upon solublisation of the membrane fraction of the cell, inclusion bodies are the densest remaining component. Upon ultracentrifugation therefore, the pellet is formed from such. The inclusion-body proteins are then solublised in a proprietary agent, with purification on protein columns.
Practice
The protocol used is identical to that given on the instructions with the kit. The 'basic' protocol is used since the pI of reflectin is 8.5.
It was found that the centrifugation forces given in the protocol for the benchtop centrifugation steps were insufficient. The speed needed to be raised by a factor of four.
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