植物激活蛋白Ap36在苏云金芽胞杆菌的表达及抗病作用

以含有植物激活蛋白(Activator protein)基因ap36的质粒pMYE为摸板，设计一对特异引物，通过PCR反应扩增出激活蛋白基因开放读码框序列，使用XbaI和SphI双切PCR产物，插入到用这两酶双切的pBMB982-304质粒slh motif的下游，构建重组表达质粒pBMB0588，转化苏云金芽胞杆菌无质粒突变株BMB171。结果表明，目标蛋白以融合蛋白的形式成功表达。为探讨表达植物激活蛋白的重组菌是否能够诱导植物获得抗性，用重组菌在28℃培养24小时发酵液原液处理番茄幼苗和土豆片。结果显示，发酵液处理90min后，番茄叶片过氧化物酶(POD)和脯氨酸含量较对照分别提高了57.14%和131.59%；4d后苯丙氨酸解氨酶(PAL)含量较对照也有所提高；经重组菌发酵液处理的马铃薯片，对马铃薯软腐病表现出很强的抗病性。

Expressing Activator Protein Ap36 in Bacillus thuringiensis and the Function of Recombined Strain on Disease Resistance

The open read frame of ap36 gene was amplified from plasmid pMYE by PCR using a pair of special primers. The PCR production was cut by XbaI and SphI and sub-cloned into the pBMB982-304. Thus, the ap36 gene sequence located the downstream of slh motif of S-layer protein gene. Then the recombinant plasmid pBMB0588 was transformed into Bacillus thuringiensis plasmid–free derivative strain BMB171. The result showed the target protein was successful expressed as a SLH-Ap36 fusion protein. In order to investigate the induced ability of the recombinant strain, tomato and potato chips treated with the 24h cultured of recombinant strain were investigated to determine the induction of defense responses. The result showed the activity of peroxidase and the amount of proline of tomato leaves was increased 57.14% and 131.59% compared to control after treatment of 90 min, respectively. The activity of phenylalanine ammonia lyase was also higher than control after treatment of 4d. Furthermore, the treated potato showed high resistance to rot disease caused by Erwinia corotovora SCG1 compared to control.