AT-rich sequences from the arbuscular mycorrhizal fungus Gigaspora rosea exhibit ARS function in the yeast Saccharomyces cerevisiae.
Autonomous replicating sequences are DNA elements that trigger DNA replication and are widely used in the development of episomal transformation vectors for fungi. In this paper, a genomic library from the mycorrhizal fungus Gigaspora rosea was constructed in the integrative plasmid YIp5 and screened in the budding yeast Saccharomyces cerevisiae for sequences that act as ARS and trigger plasmid replication. Two genetic elements (GrARS2, GrARS6) promoted high-rates of yeast transformation. Sequence analysis of these elements shows them to be AT-rich (72-80%) and to contain multiple near-matches to the yeast autonomous consensus sequences ACS and EACS. GrARS2 contained a putative miniature inverted-repeat transposable element (MITE) delimited by 28-bp terminal inverted repeats (TIRs). Disruption of this element and removal of one TIR increased plasmid stability several fold. The potential for palindromes to affect DNA replication is discussed.
Bergero
Roberta
R
Crop and Soil Research Group, Scottish Agricultural College, King's Buildings, West Mains Road, Edinburgh EH9 3JG, UK. r.bergero@ed.ac.uk
D020856:AT Rich Sequence / Q000235:genetics / Q000502:physiology*; D016384:Consensus Sequence; D004261:DNA Replication; D004251:DNA Transposable Elements / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology*; D015723:Gene Library; D038821:Mycorrhizae / Q000235:genetics*; D010957:Plasmids / Q000096:biosynthesis / Q000235:genetics; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid; D014170:Transformation, Genetic
D004251:DNA Transposable Elements; D004271:DNA, Fungal