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(12) United States Patent (10) Patent No.: US 9,394,571 B2 Ramseier Et Al

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(12) United States Patent (10) Patent No.: US 9,394,571 B2 Ramseier Et Al US0093.94571B2 (12) United States Patent (10) Patent No.: US 9,394,571 B2 Ramseier et al. (45) Date of Patent: Jul.19, 2016 (54) METHOD FOR RAPIDLY SCREENING 5,043,430 A 8, 1991 Yoshikawa MICROBAL, HOSTS TO DENTIFY CERTAIN 5,055,294 A 19 1991 Gilroy 1 STRANS WITH IMPROVEDYELD AND/OR 392. A 192: Erystal QUALITY IN THE EXPRESSION OF 5,085,862. A 2, 1992 Kleinet al. HETEROLOGOUS PROTENS 5,128,130 A 7/1992 Gilroy et al. 5,151,350 A 9, 1992 Colbert et al. (75) Inventors: Thomas M. Ramseier, Carmel, IN (US); 3.3% A 3. 3: St. Russel J. Coleman, San Diego, CA 5,169,772 A 12/1992 ZimmermanCOX et al. (US); Jane C. Schneider, San Diego, 5,173,616 A 12/1992 Hinooka CA (US); Charles D. Hershberger, 5,232,840 A 8, 1993 Olins Fremont, CA (US); Diane M. Retallack, 5,264.365 A 1 1/1993 Georgiou et al. Poway, CA (US); Charles H. Squires, 5,281,532 A 1/1994 Rammler et al. Poway, CA (US) 5,292,507 A 3, 1994 Charley s 5,292,658 A 3, 1994 Cormier et al. 5,348,867 A 9/1994 Georgiou et al. (73) Assignee: PFENEX INC., San Diego, CA (US) 5,399,684. A 3/1995 Davie et al. 5,418, 155 A 5/1995 Cormier et al. (*) Notice: Subject to any disclaimer, the term of this 5,441,934 A 8/1995 Krapcho et al. patent is extended or adjusted under 35 (Continued) U.S.C. 154(b) by 475 days. FOREIGN PATENT DOCUMENTS (21) Appl. No.: 12/109,554 EP O121352 10, 1984 (22) Filed: Apr. 25, 2008 EP O1551.89 9, 1985 (Continued) (65) Prior Publication Data OTHER PUBLICATIONS US 2008/O26907OA1 Oct. 30, 2008 Wu et al., 2002, Cell-biological applications of transfected-cell Related U.S. Application Data microarrays, Trends in Cell Biology, 12(10): 485-488.* Baneyx. F., and G. Georgiou, "Construction and Characterization of (60) signal application No. 60/914.361, filed on Apr. 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CPC CI2O I/689 (2013.01); C12N 1/20 (2013.01); Primary Examiner —Yong Pak CI2N 15/78 (2013.01) (74) Attorney, Agent, or Firm — Wilson Sonsini Goodrich & (58) Field of Classification Search Rosati CPC ........ C12N 1/20; C12N 15/78; C12Q 1 Fé89 (57) ABSTRACT See application file for complete search history. The present invention provides an array for rapidly identify (56) References Cited ing a host cell population capable of producing a heterolo gous protein with improved yield and/or quality. The array U.S. PATENT DOCUMENTS comprises one or more host cell populations that have been genetically modified to increase the expression of one or more 3,844,893 A 10, 1974 Hitzman target genes involved in protein production, decrease the 3,878,093 A 4, 1975 Kanani et al. 4,169,010 A 9, 1979 Marwill expression of one or more target genes involved in protein 4,432,895 A 2f1984 Tamowski degradation, or both. One or more of the strains in the array 4,511,503 A 4, 1985 Olson et al. may express the heterologous protein of interest in a peri 4,551,433 A 11, 1985 DeBoer plasm compartment or may secrete the heterologous protein 4,595,658 A 6, 1986 Zinder et al. extracellularly through an outer cell wall. The strain arrays 4,637,980 A 1, 1987 Auerbach et al. are useful for Screening for improved expression of any pro 4,680,264 A 7, 1987 Puhler et al. tein of interest including therapeutic proteins, hormones, 4,695.455 A 9, 1987 Bames et al. 4,755.465 A 7/1988 Gray et al. growth factors, extracellular receptors or ligands, proteases, 4,861,595 A 8, 1989 Bames et al. kinases, blood proteins, chemokines, cytokines, antibodies 4,888,274 A 12/1989 Radding et al. and the like. 4,963,495 A 10/1990 Chang et al. 5,023,171 A 6, 1991 Ho et al. 13 Claims, 3 Drawing Sheets US 9,394,571 B2 Page 2 (56) References Cited 6,077,707 6, 2000 Tsien et al. 6,080,576 6, 2000 Zambrowicz et al. 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