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Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds
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DNA primase activity from wheat embryos.

J Graveline1, L Tarrago-Litvak, M Castroviejo

  • 1Institut de Biochimie Cellulaire et Neurochimie du CNRS, 1, rue Camille Saint Saëns, 33077, Bordeaux Cedex, France.

Plant Molecular Biology
|December 7, 2013
PubMed
Summary
This summary is machine-generated.

Wheat embryo DNA primase synthesizes short RNA primers essential for DNA replication initiation. This enzyme, associated with DNA polymerase A, forms a ribo-deoxynucleotide linkage, crucial for DNA synthesis.

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Area of Science:

  • Molecular Biology
  • Enzymology

Background:

  • DNA primase is a critical enzyme for initiating DNA replication by synthesizing RNA primers.
  • Understanding plant DNA primase function is essential for comprehending DNA replication mechanisms in plants.

Purpose of the Study:

  • To characterize the DNA primase activity in wheat embryos.
  • To investigate the association of DNA primase with DNA polymerase activity.

Main Methods:

  • Enzyme purification from wheat germ and embryos.
  • Assaying DNA synthesis using various templates (poly dT, M13 ssDNA).
  • Characterization of enzyme properties (molecular weight, sedimentation coefficient) and inhibitor effects.
  • Analysis of primer synthesis and linkage formation using radiolabeled nucleotides.
  • Phosphocellulose chromatography to separate primase and polymerase activities.

Main Results:

  • Wheat embryo preparations exhibit DNA primase activity, dependent on template and enzyme.
  • The synthesized product contains a ribo-deoxynucleotide linkage, with primer size ranging from 2 to 15 residues.
  • DNA primase has a molecular weight of ~110,000 and a sedimentation coefficient of 5S.
  • Primase activity is resistant to α-amanitin and aphidicolin.
  • Phosphocellulose chromatography revealed an association between DNA primase and DNA polymerase A (γ-like).
  • DNA polymerase A specifically stimulated DNA synthesis in the presence of primase activity.

Conclusions:

  • Wheat DNA primase synthesizes RNA primers with a ribo-deoxynucleotide linkage, initiating DNA replication.
  • A DNA polymerase A (γ-like) is associated with wheat DNA primase, suggesting a functional complex.
  • This finding provides insights into the coordinated action of primase and polymerase in plant DNA replication.