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A Robotic Platform for High-throughput Protoplast Isolation and Transformation
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Colony Formation from Protoplasts of Nitrate Reductase-deficient Rice Cell Lines.

K Wakasa1, M Kobayashi, H Kamada

  • 1Department of Cell Biology, National Institute of Agrobiological Resources, Yatabe, Tsukuba, Ibaraki 305, Japan.

Journal of Plant Physiology
|December 1, 2012
PubMed
Summary

Researchers developed nitrate reductase-deficient rice cell lines using anther culture and chlorate selection. These cell lines exhibit no nitrate reductase activity and are suitable for genetic manipulation in rice.

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Area of Science:

  • Plant Biotechnology
  • Molecular Biology
  • Genetics

Background:

  • Nitrate reductase is crucial for nitrogen assimilation in plants.
  • Developing genetically modified crops requires efficient methods for generating cell lines with specific traits.

Purpose of the Study:

  • To select and characterize nitrate reductase-deficient rice cell lines.
  • To assess the potential of these cell lines for gene manipulation.

Main Methods:

  • Rice calli were cultured from microspores via anther culture.
  • Selection for nitrate reductase deficiency was performed using chlorate medium.
  • Resistant cell lines were further screened on nitrate-containing medium.
  • Enzyme activity (nitrate reductase, xanthine dehydrogenase, cytochrome c reductase) was assayed.

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  • Protoplast isolation and colony formation were evaluated.

    • Main Results:

    • Two stable nitrate reductase-deficient rice cell lines (120-2 and 144) were identified.
    • These cell lines showed no nitrate reductase activity but retained xanthine dehydrogenase activity.
    • The cell lines consisted of small aggregates, yielded viable protoplasts, and formed colonies on a modified medium.
    • Cytochrome c reductase activity was absent in the selected lines.

    Conclusions:

    • Novel nitrate reductase-deficient rice cell lines have been successfully generated.
    • These cell lines possess desirable characteristics for protoplast culture and genetic engineering.
    • The developed cell lines offer a valuable tool for future gene manipulation studies in rice.