Vectors and conditions which allow genetic transformation of cephalosporium
AssigneeEli Lilly and Company
InventorsJerry L. Chapman, Jr., Thomas D. Ingolia, Kevin R. Kaster, Stephen W. Queener, Paul L. Skatrud
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Abstract
A method for transforming Cephalosporium and other lower eukaryotes is disclosed. The method involves inserting a recombinant DNA cloning vector comprising a Saccharomyces cerevisiae transcriptional and translational activating sequence positioned for expression of hygromycin phosphotransferase into a host cell and then growing the host cell under selective conditions. The vectors optionally further comprise Cephalosporium ribosomal DNA and also sequences that allow for replication and selection in E. coli and Streptomyces.
Claims43
A method for transforming a Cephalosporium host cell which comprises:
- (1) introducing a recombinant DNA cloning vector into a Cephalosporium host cell, said vector comprising
- (a) a Saccharomyces cerevisiae transcriptional and translational activating sequence positioned for expression of a sequence that codes for hygromycin B phosphotransferase and
- (b) a sequence that codes for hygromycin B phosphotransferase, subject to the limitation that the sequence of (a) is positioned for the expression of the sequence of (b), and
- (2) growing said host cell under selective conditions suitable for maintaining DNA comprising said vector in said host cell.
A recombinant DNA cloning vector which is plasmid pPS5.
A recombinant DNA cloning vector which is selected from the group consisting of plasmids pIT221 and pPS6.
A recombinant DNA cloning vector selected from the group consisting of plasmids pIT220, pETS702 and pPLS221.
A host cell selected from the group consisting of Streptomyces lividans/pETS702, Streptomyces lividans/pPLS221, E. coli K12/pETS702, E. coli K12/pPLS221, E. coli K12 RR1/pIT221, E. coli K12 RR1/pPSI, E. coli K-12 RR1/pPS5 and E. coli K12 RR1/pPS6.
The method of claim I wherein the host cell is selected from the group consisting of Cephalosporium salmyosynnematum, C. chrysogenum, C. purpurascens and C. curtipis.
Dependencies are inferred from the claim text.