Eukaryotic DNA segments capable of autonomous replication in yeast.

Area of Science:

• Molecular Biology
• Genetics
• Yeast Biology

Background:

• Autonomous replication is crucial for DNA stability and propagation in eukaryotic cells.
• Identifying origins of replication is key to understanding DNA replication control.
• The yeast Saccharomyces cerevisiae is a model organism for studying eukaryotic DNA replication.

Purpose of the Study:

• To develop a selective scheme for isolating DNA sequences that replicate autonomously in yeast.
• To identify and characterize eukaryotic DNA sequences with replication origins.
• To explore the potential of these sequences in studying DNA replication and transformation.

Main Methods:

• Utilizing a yeast vector (YIp5) containing the ura3 gene.
• Creating molecular hybrids between YIp5 and DNA from various eukaryotes.
• Transforming a ura3 deletion mutant yeast strain with these hybrids.
• Selecting for Ura+ transformants and analyzing their growth and stability.

Main Results:

• Hybrid molecules containing eukaryotic DNA successfully transformed the ura3 deletion mutant yeast.
• Transformants exhibited slow growth and instability under nonselective conditions.
• The transforming DNA existed as autonomously replicating, supercoiled circular molecules.
• The phenotype of transformants suggests the presence of eukaryotic origins of replication.

Conclusions:

• The selective scheme effectively isolated eukaryotic DNA sequences capable of autonomous replication in yeast.
• These identified sequences likely contain functional eukaryotic origins of replication.
• The isolated sequences offer valuable tools for studying DNA replication control in yeast and other eukaryotes.