Patent ID: 7527927

Claim:
A method of integrating multiple copies of exogenous DNA into reiterated chromosomal DNA of cells, comprising: (a) transforming the cells with a replicative and integrative plasmid comprising an autonomous replicating sequence, exogenous DNA, and a first selection marker; and (b) repeatedly replicating the cells from step (a) to produce a number of generations of progeny cells while selecting for cells which include the selection marker, promoting the retention of the replicative and integrative plasmid in subsequent generations of the progeny cells to produce progeny cells having multiple integrated copies of the exogenous DNA, and wherein the method further includes the step of repeatedly replicating the progeny cells to produce a number of generations of progeny cells in the absence of selection for cells which include the selection marker, so as to promote the loss of the plasmid in subsequent generations of progeny cells and recovering the cells each containing multiple copies of the exogenous DNA integrated into its chromosomal DNA, and wherein the cells are yeast cells and the exogenous DNA includes genes encoding xylose reductase, xylitol dehydrogenase, and xylulokinase, which also serve as the first selection marker.