"System 48" high-throughput cloning and protein expression analysis

Abstract

We describe a plate-based cloning and expression strategy for efficient high-throughput generation of validated expression clones in Escherichia coli. The process incorporates 48- or 96-well plates at all stages including the cloning and colony selection phases which are often performed manually. A 48-grid agar growth plate has been integrated into the colony selection component to improve throughput at the cloning stage. The combinations of 48- and 96-well plate formats are compatible with automated liquid handlers and multichannel pipettes. This revised cloning and expression pipeline increases throughput significantly, and also results in a reduction in both time and material requirements. The system has been validated by the production and screening of several thousand clones at the Midwest Center for Structural Genomics. © 2009 Humana Press.