Ningxia Wolfberry The Establishment Of Cell Lines And In Vitro Screening Resistant Cell Variants Of Hydroxyproline (hyp) Department

Lycium barbarum L. is a perennial deciduous bush in the solanaceous family, and is an important tree for commercial fruit growing in Ningxia areas. With the further development for medical use and health care recently, the cultivated areas have increased continuously. The conventional breeding can not meet the requirements for Lycium barbarum L. improvement of varieties. The techniques, developed in 1970, for plant cell and tissue culture provide the selection of resistant varieties with an advanced method. The range and frequency of somaclonal variation will be extended widely by combining these techniques with treatment of rational factors. The application of isolating the mutant became a highlight increasingly because of its easy operation and control, lower costs and short period of breeding. According to this, we set up the system for a suspension cell line by inducing the embryonic calli from the mature embryo, isolate single cell, screen the cell line resistant to hydroxyproline (HYP) and try to provide technology and method for breeding. In this study, the three-year-old trees of Lycium barbarum L. CV. Ningqi NO. I were used as plant materials. The 18-day-old mature embryos from the fruit pollinated were taken out as explants, and inoculated in different media. Our main concerns are:( 1) effects of some factors on the cell line development using cell suspension culture: a. observations of the morphological differentiation of the embryos as explant. b. effects of different hormone concentration on the callus induction. c. effects of different time and temperature on subculture of cell suspension culture.(2) effects of some factors on plantlet regeneration from a single cell, a. studies on embryogenesis development. b. effects of different hormone concentrations on induction of embryonic calli. c. effects of different hormone concentrations on produced embryoid and its germination. d. effects of different hormone concentrations on the single cell plantlet regeneration. And then, the follow researches were done using induced embryonic calli as materials: (1) effects of the treatment with 60Co- i -ray radiation on the growth of calli. (2) selection of Hyp-resistant variants. (3) characterization and stability of HYP-tolerance variants resistant to NaC1. (4) physiological and biochemical assay of the HYP-resistant variants. (5) the differentiation of variants and physiological and biochemical analysis of regenerated plantlets. The main results are as follows: 1. The stable cell lines by the cell suspension culture are established using the induced embryonic calli derived from mature embryo. The regenerated plantlets from single cells are produced, and 14 plantlets transferred to the field are obtained. 2. A hydroxyproline (HYP) resistant cell lines of Lycium barbarum L. are isolated under selected pressure of 16 mmol/L HYP after 3OGy 60Co- rays treatment. Comparison of the free amino acid levels in the cell line with that of the donor showed substantial accumulation, of proline(3 .3 X). The free proline contents of the plantlets leaves from variant lines are 13.5 time higher than that the control's under normal condition.