Method for rapidly screening microbial hosts to identify certain strains with improved yield and/or quality in the expression of heterologous proteins
First Claim
1. A method for selecting at least one optimal expression system for use in the expression of at least one heterologous recombinant protein, the method comprising:
- a. obtaining an array assembled using a method comprising;
placing in separate addressable locations on the array at least 10 nonidentical test expression systems, said at least 10 nonidentical test expression systems each comprising a different combination ofi. a Pseudomonad or E. coli host cell population, andii. at least one expression vector encoding the at least one heterologous recombinant protein,wherein the array includes at least 5 different host cell populations and at least 2 different expression vectors, and further wherein at least 3 of said at least 5 different host cell populations are deficient in their expression of at least one protease;
b. simultaneously screening the at least 10 nonidentical test expression systems on the array, wherein at least one of the nonidentical test expression systems overexpresses the heterologous recombinant protein as compared to an indicator strain; and
c. selecting at least one optimal expression system based on improved expression of each at least one heterologous recombinant protein in the optimal expression system.
5 Assignments
0 Petitions
Accused Products
Abstract
The present invention provides an array for rapidly identifying a host cell population capable of producing a heterologous recombinant protein with improved yield and/or quality. The array comprises one or more host cell populations that have been genetically modified to increase the expression of one or more target genes involved in protein production, decrease the expression of one or more target genes involved in protein degradation, or both. One or more of the strains in the array may express the heterologous recombinant protein of interest in a periplasm compartment or may secrete the heterologous recombinant protein extracellularly through an outer cell wall. The strain arrays are useful for screening for improved expression of any protein of interest including therapeutic proteins, hormones, growth factors, extracellular receptors or ligands, proteases, kinases, blood proteins, chemokines, cytokines, antibodies and the like.
298 Citations
35 Claims
-
1. A method for selecting at least one optimal expression system for use in the expression of at least one heterologous recombinant protein, the method comprising:
-
a. obtaining an array assembled using a method comprising; placing in separate addressable locations on the array at least 10 nonidentical test expression systems, said at least 10 nonidentical test expression systems each comprising a different combination of i. a Pseudomonad or E. coli host cell population, and ii. at least one expression vector encoding the at least one heterologous recombinant protein, wherein the array includes at least 5 different host cell populations and at least 2 different expression vectors, and further wherein at least 3 of said at least 5 different host cell populations are deficient in their expression of at least one protease; b. simultaneously screening the at least 10 nonidentical test expression systems on the array, wherein at least one of the nonidentical test expression systems overexpresses the heterologous recombinant protein as compared to an indicator strain; and c. selecting at least one optimal expression system based on improved expression of each at least one heterologous recombinant protein in the optimal expression system. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35)
-
Specification