Protocol development for somatic embryogenesis, SSR markers and genetic modification of Stipagrostis pennata (Trin.) De Winter

Abstract

Background Stipagrostis pennata (Trin.) De Winter is an important species for fixing sand in shifting and semi-fixed sandy lands, for grazing, and potentially as a source of lignocellulose fibres for pulp and paper industry. The seeds have low viability, which limits uses for revegetation. Somatic embryogenesis offers an alternative method for obtaining large numbers of plants from limited seed sources. Results A protocol for plant regeneration from somatic embryos of S. pennata was developed. Somatic embryogenesis was induced on Murashige & Skoog (MS) medium supplemented with 3 mg·L–1 2,4-D subsequently shoots were induced on MS medium and supplemented with 5 mg·L–1 zeatin riboside. The highest shoots induction was obtained when embryogenic callus derived from mature embryos (96%) in combination with MS filter-sterilized medium was used from Khuzestan location. The genetic stability of regenerated plants was analysed using ten simple sequence repeats (SSR) markers from S. pennata which showed no somaclonal variation in regenerated plants from somatic embryos of S. pennata. The regenerated plants of S. pennata showed genetic stability without any somaclonal variation for the four pairs of primers that gave the expected amplicon sizes. This data seems very reliable as three of the PCR products belonged to the coding region of the genome. Furthermore, stable expression of GUS was obtained after Agrobacterium-mediated transformation using a super binary vector carried by a bacterial strain LBA4404. Conclusion To our knowledge, the current work is the first attempt to develop an in vitro protocol for somatic embryogenesis including the SSR marker analyses of regenerated plants, and Agrobacterium-mediated transformation of S. pennata that can be used for its large-scale production for commercial purposes.

Molecular and histological validation of modified in ovulo nucellus culture based high-competency direct somatic embryogenesis and amplitude true-to-the-type plantlet recovery in Kinnow mandarin

“…However, Corredoira et al (2019), in their review on the establishment of emblings, speculated the involvement of germination medium. Preconditioning of somatic embryo germinated emblings on a hormone-free medium before transferring to a hardening medium has recently been emphasized by Asadi-Aghbolaghi et al (2021). Our results also revealed a positive effect on growth and development of a liquid culture medium that suggests its superiority for better nutrient utilization, ease of plant removal, lesser damage to the roots, and solubilization of synthesized inhibitors near the root zone, thus resulting in the better establishment of the emblings.…”

“…(2019) , in their review on the establishment of emblings, speculated the involvement of germination medium. Preconditioning of somatic embryo germinated emblings on a hormone-free medium before transferring to a hardening medium has recently been emphasized by Asadi-Aghbolaghi et al. (2021) .…”

Genetic Evaluation of In Vitro Micropropagated and Regenerated Plants of Cannabis sativa L. Using SSR Molecular Markers

“…In another species of Malus , Malus niedzwetzkyana , Nurtaza et al [ 97 ] using simple sequence repeat primers, detected no somaclonal variation between the mother plant and the micropropagated clones of this endangered species. Asadi-Aghbolaghi et al [ 98 ] analyzing the genetic stability of regenerated Stipagrostis pennata plants using SSR markers, detected no somaclonal variation in regenerated plants from somatic embryos of S. pennata . Bandupriya et al [ 99 ] using SSR markers, stated that there was no somaclonal variation or genetic instability in plantlets that were regenerated from Cocos nucifera L. ovary explants.…”

Histological Approach to the Study of Morphogenesis in Callus Cultures In Vitro: A Review

“…Be that as it may, under adequate in vitro conditions, explant cells give rise to primary morphogenic calli. The morphological characteristics of calli that have appeared on the surfaces of various explants and are capable of morphogenesis during further in vitro culturation are quite similar in many plants; they are compact nodular structures [29][30][31][32] (Figure 1(1)).…”

“…Be that as it may, under adequate in vitro conditions, explant cells give rise to primary morphogenic calli. The morphological characteristics of calli that have appeared on the surfaces of various explants and are capable of morphogenesis during further in vitro culturation are quite similar in many plants; they are compact nodular structures [29][30][31][32] (Figure 1(1)). 2) data; developed primary morphogenic callus according to morphological (3) and histological (4) data; shoots (buds) in the morphogenic callus according to morphological (5) and histological (6) data; roots in the morphogenic callus according to morphological (7) and histological (8) data; gemmorhizogenic structures in the morphogenic callus according to morphological (9) and histological (10) data; somatic embryos in the morphogenic callus according to morphological (11) and histological (12) According to histological data, for example, the primary morphogenic callus obtained from the wheat embryo, in the initial stages of development, is represented by a system of meristematic cells of similar size [33] (Figure 1(2)).…”