Optimization of transient expression by Agrobacterium (agroinfiltration) in leaves and buds of barley (Hordeum vulgare L.)

Document Type : Research Paper

Authors

1 Assist. Prof., Dept. of Plant Bioproducts, Institute of Agricultural Biotechnology (IAB), National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran

2 Graduated M. Sc., Dept. of Plant Bioproducts, Institute of Agricultural Biotechnology (IAB), National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran

Abstract

Barley is one of the most important crops in the world and is also important in agricultural biotechnology as a recombinant protein production platform. In plants such as cereals, whose regeneration and transformation are difficult, in some cases transient gene expression is considered as a substitute for the stable method. Agrobacterium-based transient expression (agroinfiltration) is an inexpensive and quick way to study the function of genes and to evaluate the elements that affect gene expression (untranslated regions, introns, transcription factors and promoters). By using this method, it is possible to evaluate the function of gene constructs without time-consuming process of plant regeneration only within a few days. In order to optimize this method in barley, the effect of different treatments such as tissue type, plant age, leaves position, acetosyringone concentration, type of induction medium, time of assay after induction, Agrobacterium strain, OD of induction medium, type of procedure, duration and frequency of vacuum pump operation and barley genotype on transient expression of the ß-glucuronidase reporter gene in leaves were evaluated in this research. Also, the effect of type of induction medium, Agrobacterium strain and barley genotype on transient expression of the reporter gene in buds were evaluated. Transient gene expression of ß-glucuronidase were observed in all bud samples of Bahman cultivar and EC-82-6 genotype of barley which were inoculated with GV3301 strain harboring the reporter gene construct suspended in the induction medium with acetosyringone concentration of 200 uM by using insulin syringe. According to the results of this study, an effective and low-cost Agrobacterium-based transient expression system in barley is introduced.

Keywords

References

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Cereal Research
Volume 9, Issue 1
June 2019
Pages 71-81

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