High-throughput proteomics: a flexible and efficient pipeline for protein production.
Many studies that aim to characterize the proteome require the production of pure protein in a high-throughput format. We have developed a system for high-throughput subcloning, protein expression and purification that is simple, fast, and inexpensive. We utilized ligation-independent cloning with a custom-designed vector and developed an expression screen to test multiple parameters for optimal protein production in E. coli. A 96-well format purification protocol that produced microgram quantities of pure protein was also developed.
Doyle
Sharon A
SA
United States Department of Energy Joint Genome Institute, 2800 Mitchell Drive, Walnut Creek, California 94598, USA.
Murphy
Michael B
MB
Massi
Jennifer M
JM
Richardson
Paul M
PM
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000302:isolation & purification*
D011994:Recombinant Proteins