转CecropinB基因柑桔对溃疡病的离体抗性评价

In vitro Evaluation of Resistance to Citrus Canker by Transgenic Orange Plants with Antibacterial Peptide Gene Cecropin B

为评价转抗菌肽基因Cecropin B的锦橙植株对柑桔溃疡病的抗性,分别用喷雾法和针刺法进行了离体叶接种柑桔溃疡病菌试验,并用Real-time PCR和Southern blot进一步分析了Cecropin B在转基因植株中的表达、整合情况。结果显示,与喷雾法相比,针刺接种处理的离体叶片显症时间短,发病率高,便于统计分析。针刺接种6天,大部分转基因植株发病率与非转基因植株无显著差异;其中,PR8、PR11、AAT8和AAT14单株的发病率分别为67.0%、62.9%、68.1%和54.2%,显著低于非转基因植株(96.7%)。对柑桔溃疡病具有抗性的转基因植株(PR8、PR11、AAT8和AAT14)的Cecropin B基因表达量明显高于对照(非转基因植株)。PR8和AAT14植株中的Cecropin B为单拷贝,PR11和AAT8植株中的Cecropin B分别为2个拷贝和3个拷贝。转入Cecropin B可提高锦橙对柑桔溃疡病的抗性。

Resistance of transgenic Jincheng plants with Cecropin B to citrus canker caused by Xan- thomonas axonopodis pv. cirri （Xac） was investigated with the methods of in vitro leaf inoculation- by needle-puncture and spray inoculations. Expression of Cecropin B gene in transgenic plants was analyzed by Real time q-PCR and Southern blotting. The results showed that compared with spray inoculation, the needle puncture inoculation demonstrated the advantages on collecting data for sta- tistic analysis with earlier development of the disease symptom and higher value of incidence rate. After 6 days of inoculation, most of transgenic plants had no significant difference on incidence rate with nontransgenic plants. However, the incidence rates of PR8, PR11, AAT8 and AAT14 were 67.0% ,62.9%,68.10% and 54. 2%, respectively, which were significantly lower than those of non- transgenic plants （96.7%）. The expression of Cecropin B gene in PR8, PR11, AAT8 and AAT14 plants was also higher than that of control. Cecropin B gene in PR8 and AAT14 was single copy, and in PR11 and AAT8 plants was two and three copies, respectively. Above evidences indicated that transformation of Cecropin B gene to Jincheng orange increased resistance to Xac.