Haploid Culture and Double Haploid Induction in Medicago sativa L. cv. XinJiangDaYe

Abstract

Background: Alfalfa (Medicago Sativa), a perennial cross-pollinated plant, is one of the most important forage crops in the world with commercial value and ecological significance. However, due to the complexity of its genome, varietal improvement is difficult. Therefore, generating genetically homozygous materials have greater significance for breeding. In the current study, we aimed to identify the best tissue culture conditions to obtain haploid plants and double haploid plants.Methods: In this study, the haploid plants of alfalfa were obtained by combining tissue culture regeneration system with Flow cytometry. Different concentrations of colchicine were applied to the haploid plants using solid and liquid cultivation methods to determine the optimum conditions to obtain double haploid plants of Medicago Sativa L. cv. ‘XinJiangDaYe’. Result: Among the two colchicine cultivation methods tested, the doubling rate of regenerated plants obtained by liquid cultivation method was higher and the leaves developed under this system have the best doubling effect among the three explants tested. Optimal doubling conditions for alfalfa haploid (Medicago Sativa L. cv. ‘XinJiangDaYe’) were identified. The double haploid plant material generated from the current study could serve as a genetic resource for developing the hybrid combinations and for analyzing genetic linkage in alfalfa improvement programs.

Establishment of Tissue Culture Regeneration System for Medicago ruthenica L. cv. ‘Zhilixing’

“…The rooting rate of differentiated shoots on half-strength MS basal medium supplemented with 0.1 mg/L NAA was significantly higher than these on C4 and C5 (p<0.05, Table 4). Similar results have been obtained for alfalfa (Xu et al, 2021). Adding a low concentration of NAA to the rooting medium was supportive to the rooting of differentiated shoots, but excessive NAA inhibited the rooting process (Liu et al, 2016).…”

“…Addition of 2,4-D has been reported to promote the differentiation of calli and indirectly increase the differentiation rate (Mahmood and Razzaq, 2017). Appropriate levels of 6-BA have also been reported to reduce the browning rate and effectively improve the quality of calli (Xu et al, 2021).…”

“…Successful genetic transformation requires an efficient tissue culture protocol (Mehmet et al, 2021). The regeneration process from tissue culture is very complex and is affected by many factors, such as basal media, types and genotypes of explants, growth conditions and types and combinations of phytohormones (Xu et al, 2021). Currently, the tissue culture of Medicago ruthenica traditionally utilizes mature embryos, stems, leaves and hypocotyledonary axis as explants to produce calli capable of regenerating plantlets (Bian et al, 2009;Zhang et al, 2009;Jin et al, 1997;Liu et al, 1990).…”

​Establishment of a Regeneration System for Wild Alfalfa in Alxa, China

“…Establishing a stable and efficient tissue culture regeneration system is critical to enable utilisation of stress-resistant genes in wild plants and cultivation of new stress-resistant alfalfa varieties (Zhao et al, 2020). The tissue culture regeneration process is very complex and is affected by many factors, such as the basic medium, the type and genotype of explants, growth conditions and the type and combination of plant hormones used (Xu et al, 2021). Traditionally, tissue culture of Alfalfa uses mature embryos, stems, leaves and hypocotyls as explants to produce calli that can regenerate plants (Bian et al, 2009;Zhang et al, 2009;Liu et al, 1990).…”

In vitro embryo formation and plantlet regeneration from unpollinated ovaries of Lilium L

“…In this way, the plant resource can be greatly enriched. Therefore, in recent years, haploid culture has been studied increasingly in plants such as alfalfa ( Medicago sativa L.) (Xu et al., 2021), corn ( Zea mays L.) (Y. Liu et al., 2021), gerbera ( Gerbera hybrida ) (F. Li et al., 2020), balm ( Melissa officinalis L.) (Kästner et al., 2016), and carrots ( Daucus carota L.) (Shmykova et al., 2020).…”