Methods of recombinantly producing neutral protease originating from Paenibacillus polymyxa
AssigneeRoche Diagnostics Operations, Inc.
InventorsThomas Greiner-Stoeffele, Stefan Schoenert
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Abstract
The present disclosure provides the sequence of a Paenibacillus polymyxa preproenzyme which is the precursor of a neutral protease, expression thereof in a transformed host organism, and methods for production of the neutral protease, by recombinant means. Further, use of the recombinantly produced neutral protease is disclosed in the field of cell biology, particularly for the purpose of tissue dissociation. The disclosure also includes blends with other proteases. Further disclosed are nucleotide sequences encoding the neutral protease.
Claims10
A method of isolating living cells in vitro, the method comprising:
- (a) incubating a tissue source in vitro with a recombinant neutral protease consisting of the amino acid sequence from position 289 to position 592 of SEQ ID NO:5 for a time sufficient to proteolytically degrade extracellular matrix protein components of the tissue source;
- and
- (b) isolating living cells from the proteolytically degraded tissue source.
A method of isolating living cells in vitro, the method comprising:
- (a) incubating a tissue source in vitro with a protease blend for a time sufficient to proteolytically degrade extracellular matrix protein components of the tissue source, wherein the protease blend comprises a recombinant neutral protease consisting of the amino acid sequence from position 289 to position 592 of SEQ ID NO:
- 5, a collagenase, and a thermolysin;
- and
- (b) isolating living cells from the proteolytically degraded tissue source.
Dependencies are inferred from the claim text.