Improvement of Somatic Embryogenesis and Plantlet Conversion in Alfalfa (Medicago sativa)

Undergraduate #299
Discipline: Ecology Environmental and Earth Sciences
Subcategory: Plant Research
Session: 3
Room: Exhibit Hall A

Keith Morgan - Fort Valley State University
Co-Author(s): Sangra A, and Sarwan Dhir, Center for Biotechnology, 1005 State University Drive, Fort Valley State University, GA 31030

---

In vitro somatic embryogenesis is recognized as not only a method for regenerating entire plants, but also as a potential system for analyzing the regulation of gene expression, and the metabolite and morphological changes that occur during embryo development. Somatic embryogenesis represents a highly desirable in vitro propagation system since it can yield a large number of plants and can be coupled with cryopreservation, bioreactors, synthetic seed technologies, and genetic transformation to provide increased production and to subsidize scientific researches. To establish a high-frequency plantlet regeneration system of Alfalfa (Medicago sativa) in vitro, we used leaf explants as experimental material to improve the culture protocol. An embryogenic callus was induced on MS medium supplemented with low concentrations of 2,4-D. Indirect somatic embryogenesis which was more common achieved by using leaf explants on MS medium supplemented with low concentrations of 2,4-dichlorophenoxy acetic acid (2,4-D) and Kinetin (KN). After four weeks of leaf segment culture, the embryogenic cells rapidly formed into globular embryos, and on average 23 to 25 embryos per explant were observed. The careful examination of various stages (globular, heart, torpedo, cotyledonary) of the embryo was done. Plant regeneration was achieved on half-strength MS Medium without any hormones. To further improve, the experiments were performed to evaluate the effects of different types and concentrations of carbohydrate source on fresh callus weight, the number of somatic embryos per embryonic explant and percentage of globular embryos developing into cotyledonary embryos from the leaf explants. The study suggests that the lower sugar concentration enhances the increase in calli fresh weight, while the high concentration changes calli color and decreases its proliferation rate. The positive effect of different sugar concentration (3 and 5%) for both the number of somatic embryos and the increase in calli fresh weight was also observed. This regeneration system could be used for continuous regeneration of somatic embryos for production of Alfalfa in large scale and genetic transformation studies.

Funder Acknowledgement(s): [Acknowledgements: This study was carried out with the support of Evans-Allen #336013 GEOX 5218 project from the USDA National Institute of Food and Agriculture and from National Science Foundation HER/DUE #1834046 awarded to Sarwan Dhir, Ph.D., Director of the Center for Biotechnology, Fort Valley State University, Fort Valley, GA]

Faculty Advisor: SARWAN DHIR, dhirs0@fvsu.edu

Role: Research, Design experiments