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Related Experiment Videos

A stable bifunctional antisense transcript inhibiting gene expression in transgenic plants.

A J Delauney1, Z Tabaeizadeh, D P Verma

  • 1Centre for Plant Molecular Biology, Department of Biology, McGill University, Montreal, PQ, Canada H3A 1B1.

Proceedings of the National Academy of Sciences of the United States of America
|June 1, 1988
PubMed
Summary

This study demonstrates that antisense RNA can effectively silence gene expression in plants. Introducing specific antisense sequences targeting the chloramphenicol acetyltransferase (CAT) gene led to its inactivation, proving gene silencing in plants.

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Area of Science:

  • Plant Molecular Biology
  • Genetic Engineering
  • Gene Silencing

Background:

  • Constitutive expression of chloramphenicol acetyltransferase (CAT) activity was achieved in tobacco plants using a cauliflower mosaic virus 19S promoter.
  • Further transformation introduced antisense CAT sequences to investigate gene regulation.

Purpose of the Study:

  • To develop a method for targeted gene inactivation in plants using antisense RNA technology.
  • To assess the efficacy of antisense transcripts in reducing specific gene activity under selection pressure.

Main Methods:

  • Tobacco plants were transformed with a chimeric CAT gene and subsequently retransformed with vectors containing antisense CAT sequences.
  • Selection was applied using hygromycin resistance, and gene activity was monitored.

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  • RNA and DNA blot analyses were performed to confirm gene inactivation and DNA integrity.

    • Main Results:

    • Plants expressing antisense CAT sequences showed a significant loss of CAT activity, correlated with accumulated antisense transcripts.
    • Control plants without antisense sequences maintained CAT activity.
    • DNA analysis confirmed the stability of the introduced genes, ruling out DNA instability as the cause of inactivation.

    Conclusions:

    • Antisense RNA is an effective tool for targeted gene silencing in plants.
    • Bifunctional antisense transcripts can stably inhibit the activity of endogenous genes.
    • This approach provides a method for selecting plants with reduced target gene expression.