Tung Oil Tree Establishment Of Suspension Cell Lines And The Isolation Cloning Of Homologous Gene FLC

Vernicia fordii is an endemic to China with the characteristics of long cultivated history, many varieties and wide distribution, which is one of the four major woody oil plants in China also including Camellia oleifera, Walnut and Sapium sebiferum. The tung oil with high economic value has been playing a very important role in industry. In this article the year tung oil tree with special early-flowering character has been chosen as the research object, the plant cell suspension cultivated system is initially established, and separation and cloning of FLC homologous gene is also studied simultaneously. The main results are shown as follows:1,The callus induction was carried out with the leaves of Tungoil Tree, Based on the different explants, culture medium, as well as the type and amount of growth regulator, and the optimal hormone proportion in culture medium were screened out, The results showed that 2/3 MS+KT 0.5 (mg/ L)+2,4-D 0.3 (mg/L) were the best medium to quickly induce the better callus which had loose structure and high degree of dedifferentiation. This stydy had laid the foundation to establish cell suspension system for the next step.2,By observing the impact of different cell initiation thickness over Tungoil Tree suspension cell growth situation, a lower cell growth was found while thickness was very small. This indicated that the differentiation ability is weaker, and the cell was very difficult to breed. Undifferently, if the thickness was excessive, the growth period get shorten, and the cell life rates of later stage were low. In order to keep the stability of Tungoil Tree suspension cell department, the proper inoculated density was 1.5×104～1.0×105 mL.3,Observing growth law of Tungoil Tree suspension cell system, growth curve and theoretic growth models of Tungoil Tree suspension cell was drawing while taking wet weigh for index. The result indicated that the growth development of Tungoil Tree suspension cell can be devided into four periods including delayed phase, logarithmic phase, stationary phase and decline phase. The growth curve showed s-shaped basically.4,The system of Tungoil Tree cell suspension culture was established preliminarily:2/3 MS liquid culture medium,2,4-D 0.3 mg/L, KT 0.5 mg/L,3% sucrose, pH 5.8, diffused light or the dark raise room in 25℃, the maneuver type shake cultivation, the rotational speed are 110 rpm, every other 9d Changeed culture medium 1 time in the initial period of suspension culture were the optimal conditions. Transfer in the fresh culture medium every other 6d after 27d. 5,Taking the year tung oil tree as the material, distinctive primers was designed according to extract genome team total DNA and flower bud total RNA with the purpose to increase the partial fragments of FLC gene over Tungoil Tree using PCR. After finding the similarity on-line by Blast, the sepuence results showed lower similarity, which can not confirm the part array was a FLC gene.