| 摘要: |
| 【目的】利用根癌农杆菌(Agrobacterium tumefaciens,GV3101)介导法获得转芪合酶基因烟草，改良烟草对根黑腐病的抗性。【方法】以普通烟草品种“97204”叶片为受体，采用根癌农杆菌介导法，将芪合酶基因导入烟草基因组并进行分子检测，利用灌根法对4株转芪合酶基因烟草植株接种烟草根黑腐病菌，观察烟草地上部分和根部的变化，并测定苯丙氨酸解氨酶（PAL）、多酚氧化酶（PPO）活性。【结果】通过根癌农杆菌介导、抗性选择和分子检测，获得32株转芪合酶基因烟草。从中选取长势一致的4株转基因烟草用于抗病性鉴定，结果表明，2株在灌根接种后生长正常，另外2株在接种第7天时出现轻微黄化，3株根部无发病症状，另外1株在接种16 d后根部出现发黑症状，而对照植株接种后第3天叶片就出现黄化及萎蔫症状，随着接种时间的延长，叶片黄化加剧，到第7天时已出现明显的发病症状，15 d植株几乎死亡；转基因烟草PAL及PPO活性本底均显著高于对照，PAL活性在接种后4～10 d保持在一个较高的水平，10 d后又迅速下降，PPO活性在接种2 d后迅速升高，在接种6 d达到峰值，之后开始缓慢下降，而非转基因烟草PAL、PPO活性与转基因烟草相似，但变化幅度较小。【结论】用于抗性鉴定的4株转芪合酶基因烟草中，有3株对烟草根黑腐病的抗性较好。 |
| 关键词: 烟草 芪合酶基因 遗传转化 根黑腐病 抗病性 |
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| 基金项目:国家转基因生物新品种培育项目（2008ZX08002003，2009ZX08002008B）；陕西省科技攻关项目（2011K02-01） |
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| Genetic transformation of stilbene synthase gene and research on resistance of transgenetic tobacco for Thielaviopsis basicola disease |
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| Abstract: |
| 【Objective】The study was to obtain transgenetic tobacco with the stilbene synthase gene by Agrobacterium mediated to improve the resistance of tobacco for Thielaviopsis basicola.【Method】With the leaves of ordinary tobacco varieties “97204” as recepter,the stilbene synthase gene was transplantend into tobacco genome by Agrobacterium(Agrobacterium tumefaciens,GV3101) mediated method for molecule detection,the change of soil and root was observed and the enzymatic activity of PAL,PPO determined by irrigating root method after inoculating by T.basicola for 4 plants of transgenetic tobacco with the stilbene synthase gene.【Result】By Agrobacterium mediated,resistance choice and molecule detection,32 transgenetic tobacco with the stilbene synthase gene was obtained.Among the 4 transgenetic tobacco plants,two used for resistance research grew normally,the other 2 slightly yellowed on the seventh day.The roots of 3 transgenetic tobacco had no signs of desease,root of another transgenetic tobacco had black coating after 16 d.The plants of constract appeared and wilting signs appeared on the third day.As time extended the plant of contrstract aggravated,and it showed obvious desease signs on the seventh day.The plant almost died on the fifteeth day.The enzymatic activity of transgenetic tobacco of bottom PAL,PPO was significantly higher than constract.After irrigating root vaccination,PAL enzymatic activity of transgenetic tobacco kept a higher enzymatic activity level in 4-10 d,10 d enzymatic activity fell rapidly,although the constract had similar change.The rangeability was very small;PPO enzymatic activity of transgenetic tobacco rose quickly two days after irrigating.On the sixth day it reached the peak,then fell slowly,but the PPO enzymatic activity of nontransgenetic tobacco rose quickly two days after irrigating,then slowly fell.【Conclusion】Among the 4 transgenetic tobaccos with the stilbene synthase gene used for resistance research,3 plants expressed better resistance to T.basicola. |
| Key words: tobacco stilbene synthase gene genetic transformation Thielaviopsis basicola resistance research |
