Abstract
BackgroundDomesticated einkorn (Triticum monococcum L.) is one of the oldest cultivated cereal crops in the world. Its small genome size (~ 5.7 GB), low ploidy (2n = 2x = 14, AmAm) and high genetic polymorphism make this species very attractive for use as a diploid model for understanding the genomics and proteomics of Triticeae. Einkorn, however, is still a recalcitrant monocotyledonous species for the application of modern biotechnologies, including transgenesis. This paper reports the factors that may influence transgene delivery, integration, expression and inheritance in einkorn.ResultsIn this study, we report the successful genetic transformation of einkorn using biolistic-mediated DNA delivery. Immature embryo-derived tissues of spring einkorn were bombarded with a plasmid containing the reporter gene GFP (green fluorescent protein) driven by the rice actin promoter (act1) and the selectable bar gene (bialaphos resistance gene) driven by the maize ubiquitin promoter (ubi1). Adjustments to various parameters such as gas pressure, microcarrier size and developmental stage of target tissue were essential for successful transient and stable transformation. Bombarded einkorn tissues are recalcitrant to regenerating plants, but certain modifications of the culture medium have been shown to increase the production of transgenic events. In various experiments, independent transgenic plants were produced at frequencies ranging from 0.0 to 0.6%. Molecular analysis, marker gene expression and herbicide treatment demonstrated that gfp/bar genes were stably integrated into the einkorn genome and successfully inherited over several generations. The transgenes, as dominant loci, segregated in both Mendelian and non-Mendelian fashion due to multiple insertions. Fertile homozygous T1-T2 populations of transgenic einkorn that are resistant to herbicides were selected.ConclusionTo the best of our knowledge, this is the first report of the production of genetically modified einkorn plants. We believe that the results of our research could be a starting point for the application of the current biotechnological-based technologies, such as transgenesis and genome editing, to accelerate comparative functional genomics in einkorn and other cereals.
Highlights
Domesticated einkorn (Triticum monococcum L.) is one of the oldest cultivated cereal crops in the world
Effect of bombardment parameters on transient expression A transient Green fluorescent protein (GFP) assay was used to optimize the efficiency of plasmid delivery into einkorn cells using particle inflow gun (PIG)
The developed plants displayed both good transmission of bar/ gfp transgenes and reliable resistance to herbicide treatment. This is the first study to describe the successful production of transgenic plants in einkorn (Triticum monococcum L.), a recalcitrant diploid wheat species
Summary
Domesticated einkorn (Triticum monococcum L.) is one of the oldest cultivated cereal crops in the world. (2n = 14), is the only diploid cultivated species of wheat. Domesticated einkorn, subspecies Triticum monococcum L. ssp. It is considered one of the oldest crops, as it was domesticated around 9500 years ago, and it influenced the Neolithic Revolution [1]. Domesticated einkorn was largely used as both food and animal fodder in various areas, including the Middle East, the Balkans, the Caucasus, South Africa, and Central and Mediterranean Europe. Due to its long cultivation history in various geographical and environmental areas T. monococcum is still considered an important genomic resource for the improvement of modern wheat, especially for resistance genes against various biotic and abiotic stresses [2]. It has received interest as a health food because einkorn grains contain significantly more carotenoids, lutein and other nutritional substances than modern wheats [3]
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