【摘要】 为探讨无载体主干序列(细菌DNA)的表达框基因直接转化栽培小麦的可行性和有效性,对栽培小麦(Triticum aestivum L.)进行了bar表达框基因的基因枪转化.通过试材培养、外植体分离、bar表达框基因分离提取和纯化、基因枪轰击、诱导分化与再生培养、再生苗核酸抽提、转基因检测及除草剂涂抹试验等方法,筛选到23株全部来自幼胚愈伤的独立再生苗,其中18株显示bar表达框基因的整合,且均表现抗除草剂效应.基因型鄂麦12号的转化频率(1.55%)高于鄂恩1号(0.87%),两基因型在1100psi轰击压下的转化频率均高于900psi,其中鄂麦12号达到3.51%,明显优于鄂恩1号,且高于常规小麦基因枪的平均转化频率0.50%~1.00%.鄂麦12号幼胚在1100psi轰击压下转化bar表达框基因可获高转化效应.表明受体基因型、外植体类型、轰击压等因素在优化表达框基因转化体系中具重要作用.推断表达框基因转化方法具有片段小、环境安全、转化效率高、整合模式简单、表达高效等优势,在作物遗传转化中具有广泛应用前景.更多还原

【Abstract】 To explore the transformation efficiency on cultivated wheat with gene expression cassette lacking vector backbone sequences, transformation of wheat cultivars by biolistic bombardment with bar gene expression cassette was proceeded through isolating immature embryos and inflorescences, excising gene cassette, biolistic bombarding, regenerating, PCR amplifying and leaf-painting assaying. 23 independent plantlets from embryogenesis of immature embryos were selected, 18 of which showed the integration of bar gene expression cassette and all appeared the herbicide tolerance. The transformation frequency of genotype Emai 12 (1.55%) is higher than that of Een 1 (0.87%). The two genotypes both presented higher frequencies under 1100 psi bombarded pressure than under 900 psi, Emai 12 reached 3.51% and was higher than the general transformation frequency (0.5%-1.00%) of wheat. Genotypes, explants and bombarded pressures are vital in optimizing the transformation system of gene expression cassette. Transformation of gene expression cassette is proved to be an effective approach in genetic manipulation.更多还原