An autonomously replicating plasmid transforms Aspergillus nidulans at high frequency

Abstract

From an unstable Aspergillus nidulans colony, resulting from transformation with an A. nidulans gene bank, a plasmid was reisolated which transformed A. nidulans at a frequency of 20,000 transformants per 10(6) protoplasts at near saturation levels of transforming DNA. This represents a 250-fold enhancement of transformation efficiency over that found for typical integrative vectors such as pILJ16, the plasmid used in gene bank constructions. The plasmid, designated ARp1, is 11.5 kb in size, and consists of sequences derived from the 5.4-kb gene bank vector pILJ16, which carries the A. nidulans gene argB, and a 6.1-kb insert, designated AMA1. Southern analysis of transformant DNA showed ARp1 to be maintained in free form and not integrated into the chromosome. It has a mean copy number of 10-30 per haploid genome, and is mitotically unstable, being lost from 65% of asexual progeny of transformants. It shows similar transformational properties in A. niger and A. oryzae.