Somatic embryogenesis from in vitro anther culture of apomictic buffel grass genotypes and analysis of regenerated plants using flow cytometry

Abstract

Buffel grass is a fodder grass that reproduces mainly via apomixis. Using in vitro tissue culture in apomictic species provides alternatives that can be applied in a breeding program. The aims of this work were to establish a protocol for in vitro generation of apomictic buffel grass genotypes using anthers as explants and to evaluate the genetic stability of regenerated plants via flow cytometry (FCM). Three genotypes were able to induce embryogenic calli in Murashige and Skoog (MS) medium supplemented with 6 mg/l of 2,4-dichlorophenoxyacetic acid. Seedling regeneration occurred in a MS medium supplemented with 0.5 mg/l napthaleneacetic acid ? 1 mg/l 6-benzylaminopurin. Seedlings were derived from somatic embryos and the morphogenic process was induced using the somatic tissue of the stamens. Induction and regeneration efficiency depended on the genotype and was affected by date of tiller collection, different pretreatments or the interaction of these variables. FCM analyses in in vitro regenerated plants showed genetic instability in their nuclear DNA content. Plants with lower nuclear DNA content may indicate DNA aneuploids (8.6 %), whereas plants that had twice the value of nuclear DNA content (4.7 %) suggest in vitro polyploidization. This variation observed in apomictic genotypes provides an opportunity to obtain new variants, which may then be included as sources of genetic variability in buffel grass breeding programs.

Induction of somatic embryogenesis and evaluation of genetic stability in regenerated plants of Magnolia dealbata

“…Other studies have indicated that the genotype/species plays an important role in the frequency of genetic alterations during somatic embryogenesis (Burg et al 2007). The genotype, conditions and culture of the in vitro culture are considered to be parameters that influence genetic stability; however, some studies do not show a significant effect from these factors (Burg et al 2007, Carloni et al 2014, Mamedes-Rodrigues et al 2018. In the present study, although the explants showed different responses to the time of propagation, in general, we observed a reduction in genetic fidelity with increased cycles of induction at 2.26 μM 2,4-D (≈3 %).…”

“…Many studies on in vitro propagation have focused on optimal conditions for morphogenesis; however, conditions that are optimal for plant regeneration may not be optimal for maintenance of genetic integrity (Nehra et al 1992). Some studies conducted with SE induction have reported ≥ 90 % genetic integrity, with little influence of type and concentration of PGR or length of time in culture (Harvengt et al 2001, Lopes et al 2006, Yang et al 2008, Marum et al 2009, Carloni et al 2014, Li et al 2014, Carra et al 2016, Naz et al 2016, Niazian et al 2017, Raji et al 2018. In contrast, there are also reports of high genetic variation: in Picea mariana, a phenotypic variation of 11 -57 % was reported and was associated with genotype and length in culture (Tremblay et al 1999).…”

Optimization of somatic embryogenesis and in vitro plant regeneration of Urochloa species using picloram

“…Some cultivars of the apomictic grass Pennisetum ciliare showed recalcitrant behavior when mature embryos were used as the initial explant (Colomba et al 2006). When anthers of the same cultivars were used as explants, embryogenic calluses were observed in up to 44% of the explants, and normal plants were regenerated (Carloni et al 2014). However, the advantage of using mature seed over other explants, immature embryos, or inflorescences is that they are available irrespective to the growing season.…”

In vitro regeneration of Feizixiao litchi (Litchi chinensis Sonn.)

“…Thus, GA 3 promoted rooting, inhibited root growth, and suppressed lateral root growth. Successful germination and plant regeneration via SEs have been reported with combinations of KT and NAA in ginger and buffel grass (Lincy et al, 2009;Carloni et al, 2014). After germination, GA 3 is required for plant development in buffel grass.…”