Genetic Transformation of Apomictic Grasses: Progress and Constraints

Abstract

The available methods for plant transformation and expansion beyond its limits remain especially critical for crop improvement. For grass species, this is even more critical, mainly due to drawbacks in in vitro regeneration. Despite the existence of many protocols in grasses to achieve genetic transformation through Agrobacterium or biolistic gene delivery, their efficiencies are genotype-dependent and still very low due to the recalcitrance of these species to in vitro regeneration. Many plant transformation facilities for cereals and other important crops may be found around the world in universities and enterprises, but this is not the case for apomictic species, many of which are C4 grasses. Moreover, apomixis (asexual reproduction by seeds) represents an additional constraint for breeding. However, the transformation of an apomictic clone is an attractive strategy, as the transgene is immediately fixed in a highly adapted genetic background, capable of large-scale clonal propagation. With the exception of some species like Brachiaria brizantha which is planted in approximately 100 M ha in Brazil, apomixis is almost non-present in economically important crops. However, as it is sometimes present in their wild relatives, the main goal is to transfer this trait to crops to fix heterosis. Until now this has been a difficult task, mainly because many aspects of apomixis are unknown. Over the last few years, many candidate genes have been identified and attempts have been made to characterize them functionally in Arabidopsis and rice. However, functional analysis in true apomictic species lags far behind, mainly due to the complexity of its genomes, of the trait itself, and the lack of efficient genetic transformation protocols. In this study, we review the current status of the in vitro culture and genetic transformation methods focusing on apomictic grasses, and the prospects for the application of new tools assayed in other related species, with two aims: to pave the way for discovering the molecular pathways involved in apomixis and to develop new capacities for breeding purposes because many of these grasses are important forage or biofuel resources.

An Agrobacterium strain auxotrophic for methionine is useful for switchgrass transformation

“…As contamination is a major problem with no easy solution when explants are taken from greenhouse-grown plants, in vitro-produced aseptic plants were used to work around the issue of contamination (Leifert et al 1994 ). Importantly, the use of in vitro clonally propagated plantlets as the explant donor allows for maintenance of genotype fidelity, which, barring apomictic grass species (Bellido et al 2021 ), would be lost if seed-derived explants were used. Finally, the in vitro plants are much smaller in size than greenhouse-grown plants, and thus require less greenhouse space for plant maintenance.…”

A comprehensive review of in planta stable transformation strategies

“…Historically, the most common definitions of in planta transformation have been (i) a means of transformation without tissue culture step [5,7] and (ii) a means of transformation of intact plants or plant tissues without callus culture or regeneration [14,15]. In our opinion, these definitions are incomplete and not nuanced enough to take into account the broad diversity of available in planta methods.…”

Genome-editing opportunities to enhance cattle productivity in the tropics

“…The developing of GnEd systems in a nongenetic model plants is challenging (Shan et al 2020). Major bottlenecks for GnEd in tropical forage grasses include the large and usually polyploid genomes of these species whose sequences are not available for most of them (Simeão et al 2021), and the inexistency or inefficiency of transformation protocols for a number of these species (Bellido et al 2021). Thus, more scientific developments are needed for GnEd of tropical forage grasses be able to significantly contribute with improvements in animal production.…”