Title

Engineering cytoplasmic male sterility via the chloroplast genome by expression of beta-ketothiolase

Authors

Authors

O. N. Ruiz;H. Daniell

Comments

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Abbreviated Journal Title

Plant Physiol.

Keywords

TRANSGENIC TOBACCO PLANTS; ACETYL-COA CARBOXYLASE; GENE-EXPRESSION; MOLECULAR CHARACTERIZATION; ARABIDOPSIS-THALIANA; RESTORES FERTILITY; COENZYME-A; PROTEIN; POLYHYDROXYBUTYRATE; ACCUMULATION; Plant Sciences

Abstract

While investigating expression of the polydroxybutyrate pathway in transgenic chloroplasts, we addressed the specific role of beta-ketothiolase. Therefore, we expressed the phaA gene via the chloroplast genome. Prior attempts to express the phaA gene in transgenic plants were unsuccessful. We studied the effect of light regulation of the phaA gene using the psbA promoter and 5' untranslated region, and evaluated expression under different photoperiods. Stable transgene integration into the chloroplast genome and homoplasmy were confirmed by Southern analysis. The phaA gene was efficiently transcribed in all tissue types examined, including leaves, flowers, and anthers. Coomassie-stained gel and western blots confirmed hyperexpression of beta-ketothiolase in leaves and anthers, with proportionately high levels of enzyme activity. The transgenic lines were normal except for the male-sterile phenotype, lacking pollen. Scanning electron microscopy revealed a collapsed morphology of the pollen grains. Floral developmental studies revealed that transgenic lines showed an accelerated pattern of anther development, affecting their maturation, and resulted in aberrant tissue patterns. Abnormal thickening of the outer wall, enlarged endothecium, and vacuolation affected pollen grains and resulted in the irregular shape or collapsed phenotype. Reversibility of the male-sterile phenotype was observed under continuous illumination, resulting in viable pollen and copious amount of seeds. This study results in the first engineered cytoplasmic male-sterility system in plants, offers a new tool for transgene containment for both nuclear and organelle genomes, and provides an expedient mechanism for F-1 hybrid seed production.

Journal Title

Plant Physiology

Volume

138

Issue/Number

3

Publication Date

1-1-2005

Document Type

Article

Language

English

First Page

1232

Last Page

1246

WOS Identifier

WOS:000230414800006

ISSN

0032-0889

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