Search

A series of DNA sequences was rescued from the yeast Saccharomyces cerevisiae transformed by a gene library and selected for the cdc35ts+, TRP1+ phenotype. These sequences did not complement the cdc35ts mutation, and were found in various amounts and orientations in degraded plasmids. A similar phenomenon was demonstrated when the HIS3 gene was cloned into one of them: a highly deleted plasmid was rescued from complemented homozygous diploid yeast cells, in which the HIS3+/his3- character was inherited at a 2:2 ratio. These results suggest that the insert sequences rescued from the cdc35ts transformants stimulate vigorous non-reciprocal recombination events by the transfer of HIS3 gene or the TRP-ARS fragment. This event was detected in the transformation of cdc35 or his3- hosts and was followed by the re-isolation of the degraded plasmid molecules.

A large plasmid DNA molecule was purified from Rhizobium meliloti 41 by CsCl-ethidium bromide density gradient centrifugation. Electron microscopic and agarose gel electrophoretic data suggest that addition of alkali effectively removes the chromosomal DNA, the plasmid DNA can be precipitated from the cleared lysate and no gradient centrifugation is needed for plasmid purification.