A competence of embryo-derived tissues of tetraploid cultivated wheat species Triticum dicoccum and Triticum timopheevii for efficient and stable transgenesis mediated by particle inflow gun

Dmitry Miroshnichenko,Alexander Pushin,Sergey Dolgov,Anna Klementyeva

doi:10.1186/s12870-020-02580-4

Abstract

BackgroundThe ability to engineer cereal crops by gene transfer technology is a powerful and informative tool for discovering and studying functions of genes controlling environmental adaptability and nutritional value. Tetraploid wheat species such as emmer wheat and Timopheevi wheat are the oldest cereal crops cultivated in various world areas long before the Christian era. Nowadays, these hulled wheat species are gaining new interest as donors for gene pools responsible for the improved grain yield and quality, tolerance for abiotic and biotic stress, resistance to pests and disease. The establishing of efficient gene transfer techniques for emmer and Timopheevi wheat may help in creation of modern polyploid wheat varieties.ResultsIn the present study, we describe a robust protocol for the production of fertile transgenic plants of cultivated emmer wheat (Russian cv. ‘Runo’) using a biolistic delivery of a plasmid encoding the gene of green fluorescent protein (GFP) and an herbicide resistance gene (BAR). Both the origin of target tissues (mature or immature embryos) and the type of morphogenic calli (white or translucent) influenced the efficiency of stable transgenic plant production in emmer wheat. The bombardment of nodular white compact calluses is a major factor allowed to achieve the highest transformation efficiency of emmer wheat (on average, 12.9%) confirmed by fluorescence, PCR, and Southern blot. In the absence of donor plants for isolation of immature embryos, mature embryo-derived calluses could be used as alternative tissues for recovering transgenic emmer plants with a frequency of 2.1%. The biolistic procedure based on the bombardment of immature embryo-derived calluses was also successful for the generation of transgenic Triticum timopheevii wheat plants (transformation efficiency of 0.5%). Most of the primary events transmitted the transgene expression to the sexual progeny.ConclusionThe procedures described here can be further used to study the functional biology and contribute to the agronomic improvement of wheat. We also recommend involving in such research the Russian emmer wheat cv. ‘Runo’, which demonstrates a high capacity for biolistic-mediated transformation, exceeding the previously reported values for different genotypes of polyploid wheat.

Highlights

The ability to engineer cereal crops by gene transfer technology is a powerful and informative tool for discovering and studying functions of genes controlling environmental adaptability and nutritional value
After the transferring to differentiation medium (DM), the only such cluster could produce a series of transgenic somatic embryos (Fig. 1i), which subsequently turned into two-to-five transgenic plants on RM
We found that white nodular embryogenic/organogenic calli produce more transgenic events than softer and watered morphogenic structures

Summary

Introduction

The ability to engineer cereal crops by gene transfer technology is a powerful and informative tool for discovering and studying functions of genes controlling environmental adaptability and nutritional value Tetraploid wheat species such as emmer wheat and Timopheevi wheat are the oldest cereal crops cultivated in various world areas long before the Christian era. Araraticum) are no longer cultivated [5] Both emmer and Timopheevi wheat are self-pollinated species that are characterized by persistent enclosing hulls. In recent decades, these species have attracted considerable attention of breeders as a natural source of the gene pool for the production of new immune varieties of wheat [2, 3, 6, 7]. They used in various studies as models for studying the evolution and polyploidy of the wheat genome [3, 8, 9]

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