MICROORGANISM TRANSFORMATION

摘要

Linear transforming segments are prepared having reduced vector DNA. A vector incorporating a segment having a region of homology with a target chromosomal site of a host microorganism is severed at two sites outside of the segment to remove vector sequences and at a site within the region of homology to form two segment portions, each with a part of the region of homology at one end thereof. The segment is reassembled with one of said parts of the region of homology at each end and with the ends of the segment portions opposed to the parts of the region of homology connected together. The sequence desired for integration, such as a gene, may be incorporated in the segment either intermediate the regions of homology at the ends or they may form the regions of homology. Alternatively, the linear transforming segment is prepared by circularizing a DNA segment containing the desired sequence and having the region of homology and then amplifying this construct by inverse polymerase chain reaction. Host microorganisms are transformed with these segments to obtain transformants with reduced vector DNA and particularly amylolytic S. cerevisiae transformants incorporating glucoamylase genes of S. diastaticus.