| Hemerocallis middendorffii Trautv is a plant species in the subfamily Hemerocallidoideae of the family Xanthorrhoeaceae of the order Asparagales. It is native to the Russian Far East, northwest China, Korea, and Japan. It grows in meadows, mountain slopes, open woods, and scrub. It is cultivated in Asia for its edible flowers.The leave lenth of Hemerocallis middendorfii Trautv is 50-80 cm and the width is 1-2cm. 5–6 flowers are in a terminal head, about20–90 cm, golden yellow or clear orange. Flowers often rebloom in July for 2–3 weeks and last in September. Hemerocallis middendorfii was selected as experimental materials,which is a perennial herbaceous plant of Liliaceae Hemerocallis. ‘Early Golden Flower’ is the most cold-resistant and early blossoming of all Hemerocallis, Which has broad application prospects, and it is suitable for planting in the courtyard, square, campus, flower bed.Because of the serious soil salinization in the western region of China, the local crop growth and agricultural productivity have been restricted. Along with the development of modern biological technology, people can study it from the molecular biology level which opens up a new way to improve the tolerance ability of ornamental plants. The purpose of this experiment is to improve the selection range and color matching of flowers in the early spring of the western region, and to enlarge the planting area. Improved variety of Hemerocallis middendorffii Trautv were obtained with application of plant tissue culture and the method of agrobacterium mediated to add green plant materials for the western region.Agrobacterium Mediated was used to make Lc Chi2 genetic transformation to Hemerocallis middendorfii callus. Lc Chi2 gene belongs to Chitinase19 gene family, Class II Chitinase, which has herbicide resistance, saline-alkaline tolerance, disease resistance and other functions. Studied the effect of infection concentration of Agrobacterium and acetyl- clove ketone(AS) on the conversion rate,and antibacterial effect of Cef on engineering bacteria during the transformation process. We made Na Cl+Na2CO3 mixed salt alkaline stress to the transformation Lc Chi2 bracts Hemerocallis, and then detected soluble sugar,chlorophyll, and cell membrane permeability,which related to salt tolerance physiological indexes. This study can provide technical basis for the production and promotion of Hemerocallis middendorffii,and the research results showed that,1, The optimal explants disinfection solution of Hemerocallis middendorfii plant tissueculture was: 75% alcohol for 30 s + 0.1% Hg Cl2 for 9 minutes. The optimal medium for callus inductionwas: MS+6-BA1.39mg/L+NAA0.3mg/L+2,4-D0.3 mg/L; the optimal root ing culture mediumwas: 1/2MS sucrose 20 g +NAA0.3 mg/L+2g/L activated carbon.2,The optimal transformation acceptor was plant callus. The optimal time of infection was 10 min. T he optimum infection concentration of bacterium suspension OD600 was 1.0. The optimal acetyl clove ketone concentration(AS) was 1.5 mol / L. The antimicrobial concentration of the antibiotic was 300mg/L.400mg/L glufosinate concentration was the sensitive concentration to plants.3,The extraction methods of two kinds of plant genome DNA were compared, and the result showed that genome DNA extracted with CTAB method had high yield, high purity, more suitable for Hemerocallis middendorfii leaf DNA extraction; 10 strains were amplified a specific fragment from PCR detection.4,The content of soluble sugar increased with the increase of salt and alkali stress, and the content of chlorophyll decreased with the increase of salt concentration. Non conversion group was significantly larger than that of the transformed Lc Chi2 group; cell conductivity was measured in the two groups of plants, and non electrical conductivity was greater than that of the transformed Lc Chi2 group. |
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