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Related Experiment Videos

Sequence analysis of ARS elements in fission yeast.

K Maundrell1, A Hutchison, S Shall

  • 1School of Biological Sciences, University of Sussex, Brighton, UK.

The EMBO Journal
|July 1, 1988
PubMed
Summary

Researchers identified AT-rich sequences in yeast DNA, similar to ARS elements. These sequences contain a conserved core, suggesting a role in chromosomal function, even if not essential for plasmid maintenance.

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Area of Science:

  • Molecular Biology
  • Yeast Genetics

Background:

  • Schizosaccharomyces pombe (S. pombe) chromosomal DNA possesses sequences analogous to Autonomously Replicating Sequence (ARS) elements found in Saccharomyces cerevisiae.
  • ARS elements are crucial for the initiation of DNA replication in eukaryotic chromosomes.

Purpose of the Study:

  • To identify and characterize ARS elements in S. pombe.
  • To determine the nucleotide sequence and functional significance of these ARS elements and their associated conserved motifs.

Main Methods:

  • Sau3A fragmentation of S. pombe genomic DNA.
  • Cloning of fragments into an M13-based shuttle vector.
  • Nucleotide sequencing of ARS+ inserts.
  • Deletion mapping and in vitro mutagenesis.
  • Transformation assays to assess ARS activity.

Main Results:

  • Eight ARS+ inserts were sequenced, ranging from 0.8 to 1.8 kb.
  • All sequenced ARS+ clones were AT-rich (69-75% A+T).
  • A conserved 11 bp AT-rich core element (5' (A/T)PuTT-TATTTA(A/T) 3') was identified in all ARS+ clones.
  • Deletion mapping placed the consensus core near the functional ARS domain.
  • Precise excision of the consensus core had minimal impact on ARS activity in transformation assays.

Conclusions:

  • The conserved AT-rich core element is consistently found near the functional ARS domain in S. pombe.
  • While not essential for extrachromosomal plasmid maintenance, the consensus sequence likely plays a functional role in the native chromosomal context.
  • The consensus sequence may represent a cryptic element important for ARS function in situ.

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