CONSTRUCTION OF STABLE EPISOMES IN CRYPTOCOCCUS-NEOFORMANS

We report the generation of stable plasmids constructed by inserting s pecific DNA sequences into previously known unstable vectors. These se quences were obtained from a DNA library recovered from a previously r eported stable minichromosome created by electroporative transformatio n in Cryptococcus neoformans (Varma and Kwon-Chung 1994). A 6-kb inser t from this minichromosome significantly enhanced both the frequencies at which URA5 transformants were obtained as well as the stability of their uracil prototrophy on non-selective media. A 1.5-kb sequence of this insert contained telomeric sequence repeats which when introduce d into plasmids resulted in significant increases in transformation fr equency. A 1081-bp sequence (STAB), present in the remainder of the in sert, had an ARS-like function enhancing the episomal maintenance of p lasmids in the transformants regardless of the gene (ADE2/URA5) used a s a selection marker.