Dissertation > Excellent graduate degree dissertation topics show

Studies on Regeneration and Agrobacterium-mediated Gentic Transformation of Cucumber (Cucumis Sativus L.)

Author: ZhangShouJie
Tutor: WangXiuFeng
School: Shandong Agricultural University
Course: Olericulture
Keywords: Cucumber Cotyledonary node Agrobacterium tumefaciens Transformation Phospholipase Dαgene
CLC: S642.2
Type: Master's thesis
Year: 2011
Downloads: 78
Quote: 0
Read: Download Dissertation

Abstract

Cucumber (Cucumis sativus L.) is one of the most important vegetable crops cultured both in facilities and in the open. The production of cucumber is often subjected to stress factors such as drought, salinity, pests and other factors of stresses, which seriously influenced the quality and yield of cucumber. Gene engineering provide a novel and efficiency way which enhance the breeding technology from the macro level to microcosmic level can improve the crop quality. Using biological engineering technology to foster new cucumber cultivars in high yield, excellent resistant, good quality have become the development direction of cucumber breeding. Efficient regeneration and transformation system of cucumber is the foundation of transgenic breeding. Therefore, it has important theoretical and practical significance to research the factors that influence the regeneration and transformation of cucumber and to optimize the regeneration and transformation system.Based on previous studies, we researched the various factors which influence the regeneration of cucumber, for example, the type of explants, hormone varieties and concentration, explant cutting and inoculation method, the age of seedling explants, endogenous hormonal changes, etc. We established of a complete and efficient regeneration system of cucumber using cotyledonary node, and on this basis, we also established Agrobacterium-mediated transformation system by selection pressure test on kanamycine. Using this method, we obtained antisense cuPLDαgene transgenic cucumber plants.The main results obtained are as follows:1.Cotyledonary node is the best explants to regeneration. It was influenced by exogenous plant hormones, explant cutting and inoculation method, the age of seedling explants during cotyledonary node regeneration. MS + 1.5 mg/L BA was the best medium to induce adventitious buds guide cotyledonary nodes; removal 1/3 cotyledon distal axis and cut the cotyledon edge was the best way of cutting and inoculation; 5-6 d seedling age explants were efficiency . The best rooting medium was MS without any hormone. The whole process of regeneration need about 50 d from cotyledon inoculation to regenerated plants transplanted to soil.2.We measured the endogenous hormone in the cucumber regeneration process. The results showed that: ABA content was highest in cotyledonary nodes cultured in vitro on 0 day, IAA and GA contents were minimum; the contents of ABA, IAA, GA and ZR increased at first and then decreased in the callus induction period (3-9 d); in callus differentiation period (9-15 d), IAA and GA contents first increased and then decreased, ZR content was reduced, while ABA content first decreased and then increased. It indicated that it was important of endogenous hormone to regeneration of cucumber. The endogenous hormones contents changed with the different stage of cucumber regeneration. The types, concentration and coordination balance of endogenous hormones were pivotal in bud formation and differentiation.3.Based on the regeneration of cucumber, we established efficiency Agrobacterium-mediated transformation system, mainly including preculture, coincubate, differentiation culture, and rooting culture. The production of non-transformed adventitious buds was inhibited on 150 mg/L Kanamycine. 250 mg/L carbenicilin can effectively restrain the growth of Agrobacterium. 20 transformed plants were obtained from 250 cotyledonary node explants and 12 transgenic plants were confirmed using PCR specific analysis, and the transgenic rate was approximately 4.8 %. The expression level of cuPLDαgene in transgenic plants were markedly lower than in wildtype plants, indicating that the entire foreign gene has been successfully inserted in cucumber genome and inhibited the cuPLDαgene expression

Related Dissertations

CLC: > Agricultural Sciences > Gardening > Vegetable gardening > Melons > Cucumber
© 2012 www.DissertationTopic.Net Mobile