WHOLE CELL ENGINEERING BY MUTAGENIZING A SUBSTANTIAL PORTION OF A STARTING GENOME, COMBINING MUTATIONS, AND OPTIONALLY REPEATING

摘要

An invention comprising cellular transformation, directed evolution, and screening methods for creating novel transgenic organisms having desirable properties. In one embodiment, this invention provides a method of generating a transgenic organism, such as a microbe or a plant, having a plurality of traits that are differentially activatable. This invention also provides a method of retooling genes and gene pathways by the introduction of regulatory sequences, such as promoters, that are operable in an intended host, this conferring operability to a novel gene pathway when it is introduced into an intended host. For example a novel man-made gene pathway, generated based on microbially-derived progenitor templates, that is operable in a plant cell. This invention also provides a method of generating novel host organisms having increased expression of desirable traits, recombinant genes, and gene products. This invention provides novel methods for determining polypeptide profiles, and protein expression variations, which methods are applicable to all sample types disclosed herein. The present invention provides methods of simultaneously identifying and quantifying individual proteins in complex protein mixtures. Additionally this invention provides methods for cellular and metabolic engineering of new and modified phenotypes by using "on-line" or "real-time" metabolic flux analysis.