CAJ | 학술논문

葡萄糖氧化酶催化分子氧氧化b-D葡萄糖产生葡萄糖酸和H2O2.产生H2O2和其它活性氧物质是植物对病原体侵入的防御反应.实验证明H2O2水平升高不仅可诱发细胞过敏坏死反应,而且还能激活抗病基因表达如植物抗毒素及诱导病原相关蛋白(PR)等基因的表达.通过农杆菌介导将来自黑麴霉(Aspergillus niger)的葡萄糖氧化酶基因转入2个马铃薯重要栽培品种大西洋(Atlantic)和夏普蒂(Shepody)中获得23个转基因株系,其中75%为Kan抗性植株.在23个转基因株系中有16个株系可用PCR扩增出目的基因.进一步用核酸斑点杂交选出13个杂交阳性株系.转基因植物总DNA的Southern blot 分析表明,GO基因已整合到马铃薯四倍体基因组中,转基因大西洋植株中目的基因为2～4 个拷贝,而夏普蒂转基因植株中为1个拷贝.将转基因植株离体叶片接种呼和浩特地区流行的Phytophthoro infestans的复合生理小种1,3,4孢子.结果表明,和未转基因对照相比转基因植株病斑出现时间晚,病斑扩展速度慢,发病程度轻,感病叶片少.总体上表现出较明显抗病性.转基因大西洋中抗性株系多于夏普蒂.转基因马铃薯植株体内H2O2含量测定表明,转基因马铃薯植株的根系和叶片在KI-淀粉培养基上数天后变为蓝色,而未转基因对照植株的根系和叶片变化不明显.试验表明,转基因马铃薯植株对晚疫病的抗性和体内H2O2含量水平呈正相关.还对马铃薯转化中再生小植株诱导,转基因植株中GO基因拷贝数,对晚疫病抗性的鉴定以及表达葡萄糖氧化酶基因的转基因马铃薯产生抗性的机理等问题进行了讨论.
포도당양화매최화분자양양화b-D포도당산생포도당산화H2O2.산생H2O2화기타활성양물질시식물대병원체침입적방어반응.실험증명H2O2수평승고불부가유발세포과민배사반응,이차환능격활항병기인표체여식물항독소급유도병원상관단백(PR)등기인적표체.통과농간균개도장래자흑국매(Aspergillus niger)적포도당양화매기인전입2개마령서중요재배품충대서양(Atlantic)화하보체(Shepody)중획득23개전기인주계,기중75%위Kan항성식주.재23개전기인주계중유16개주계가용PCR확증출목적기인.진일보용핵산반점잡교선출13개잡교양성주계.전기인식물총DNA적Southern blot 분석표명,GO기인이정합도마령서사배체기인조중,전기인대서양식주중목적기인위2～4 개고패,이하보체전기인식주중위1개고패.장전기인식주리체협편접충호화호특지구류행적Phytophthoro infestans적복합생리소충1,3,4포자.결과표명,화미전기인대조상비전기인식주병반출현시간만,병반확전속도만,발병정도경,감병협편소.총체상표현출교명현항병성.전기인대서양중항성주계다우하보체.전기인마령서식주체내H2O2함량측정표명,전기인마령서식주적근계화협편재KI-정분배양기상수천후변위람색,이미전기인대조식주적근계화협편변화불명현.시험표명,전기인마령서식주대만역병적항성화체내H2O2함량수평정정상관.환대마령서전화중재생소식주유도,전기인식주중GO기인고패수,대만역병항성적감정이급표체포도당양화매기인적전기인마령서산생항성적궤리등문제진행료토론.
Two commercial potato cultivars "Atlantic" and "Shepody" were transformed with glucose oxidase gene from Aspergillus niger via Agrobacterium tumefaciens mediated gene transfer. 23 regenerated transgenic potato plants were obtained. Kanamycin resistance, PCR amplification, nucleic acid spot hybridization, southern blot analysis showed that the glucose oxidase gene was intergrated into transformed potato genome. Southern blot analysis revealed that there are 2～4 copies of glucose oxidase gene in each potato tetraploid genome. Detached transgenic potato leaves were inoculated with spors of Phytophthora infestans. Tests show that the number and the size of lesions were reduced and the appearance of lesions was postponed on the leaves of some transgenic lines.