农杆菌介导CBF1基因转化安祖花的研究

目的]为获得耐低温安祖花奠定基础。方法]利用农杆菌介导CBF1基因侵染安祖花,并研究不同浓度乙酰丁香酮(AS)和侵染时间对转化后安祖花抗性愈伤组织的存活率和抗性芽分化率的影响。结果]以重组质粒pBI121-CBF1为模板,利用CBF1cDNA全长的引物序列进行PCR扩增,获得约650bp的片段,表明重组质粒构建成功。在抗性培养基上筛选12周后,部分愈伤组织褐化死亡。用100μmol/LAS作为诱导物质时,抗性愈伤组织存活率和抗性芽的分化率较高。农杆菌侵染时间对转化后愈伤组织存活率和抗性芽分化率有一定影响。结论]当AS浓度为100μmol/L、侵染时间为10min时,抗性愈伤组织的存活率及分化率最高,分别为48.6%和11.46%。

Study on the Transformation of Anthurium andreanum by Agribacterium tumefacien -mediated CBF1 Gene

Objective] The aim of the research was to lay the foundation for obtaining Anthurium andraeanum Lind with tolerance to low temperature. Method] A. andraeanum was infected by using CBF1 gene mediated by Agrobacterium tumefacien to study the effects of different concn. of acetosyringone (AS) and different infection time on the survival rate of resistant callus and the differentiation rate of resistant buds in A. andraeanum after transformation. Result] With the recombinant plasmid pBI121-CBF1 as template, PCR amplification was made by using the whole primer sequence of cDNA of CBF1 gene and one fragment with the length of about 650 bp was obtained, which indicated that the recombinanted plasmid containing CBF1 gene was constructe successfully. After screening on the resistant medium for 12 weeks, some calli browned and died. When 100 umol/L AS was used as the inducing materials, the survival rate of resistant calli and the differentiation rate of resistant buds were higher. The infection time of A. tumefacien had certain effects on the survival rate of resistant calli and the differentiation rate of resistant buds after transformation. Conclusion] When AS concn. was 100 μmol/L and the infection time was 10 min, the survival rate of resistant calli and the differentiation rate of resistant buds were highest, being 48.65% and 11.46% resp.