【作者】 解修超；

【导师】 张荣意；

【作者基本信息】 华南热带农业大学 ， 植物病理学， 2004， 硕士

【摘要】 采用组织分离法，从4～7成熟度的香蕉果实中分离到香蕉果实炭疽菌（Colletotrichum musae）弱致病株Z₁、Z₂和强致病株Z₃、Z₄。研究发现强致病株和弱致病株的培养性状也存在明显的差别。 在香蕉果皮、叶、苞片、花器及假茎上均有香蕉炭疽病菌潜伏，假茎的病残体带菌率最高，为主要的侵染来源；采后香蕉果实上炭疽病侵染型为潜伏侵染型；开花后20d 的香蕉果实开始带菌，随时间延长，带菌量逐渐增加，以后各个生长阶段的带菌率与降雨量有关；炭疽病菌在果指不同部位的分布存在差异。 采用徒手切片法和石蜡切片法观察香蕉果实炭疽菌的侵染过程，接种在香蕉果实上的分生孢子6hr萌发，12hr形成淡色附着胞，24hr形成暗色附着胞，30d后未见附着胞形成侵入丝。暗色附着胞绝大部分潜伏在果实表皮细胞间隙，并且一直潜伏到香蕉果实采收。分生孢子在叶部、假茎与在果实上的发育过程相似，但假茎上暗色附着胞形成部位不象叶部和果实上那样局限于细胞间隙。 根据香蕉炭疽菌的两个特异片段（分别为357bp和206bp），设计两对引物：采用SDS法分别提取了香蕉组培苗基因组DNA、香蕉果实炭疽菌弱致病株Z₁基因组DNA、香蕉果实炭疽菌强致病株Z₄基因组DNA、芒果炭疽菌基因组DNA、橡胶炭疽菌基因组DNA、柱花草炭疽菌基因组DNA、西瓜炭疽菌基因组DNA、香蕉冠腐病菌基因组DNA、西瓜枯萎病菌基因组DNA；以上述基因组DNA为模板对特异片段进行PCR验证，证明357bp的片段为Colletotrichum musaes所特有，可以用此片段进行香蕉果实炭疽病的分子检测试验，。 分子检测预试验结果表明：香蕉组培苗基因组DNA对香蕉炭疽菌基因组DNA产生严重的干扰，当香蕉组培苗基因组DNA的用量为香蕉炭疽菌基因组DNA用量的2倍和2倍以上时，目标带检测不到。华南热带农业大学2004届硕七学位论文 对混合DNA进行酶切，用酶切后的产物作模板进行PCR检测，结果用EcoRI酶切后的产物作模板的PCR，，检测到了目标带，而用Pstl、Hindln的酶切后的产物作模板的PCR，未检测到目标带。 香蕉果实接种后PCR分子检测试验结果表明:但低浓度的抱子悬浮液接种香蕉果实后，未检测到目标带;香蕉果实被高浓度的抱子悬浮液接种后，检测到了目标带。更多还原

【Abstract】 The pathogen of banana fruit anthracnose was isolated from 30~70 days banana fruit using organic separation method. The isolated pathogen included hypovirulent strains Z1, Z2 and high virulent strains Z3, Z4. The cultural characteristics between hypovirulent strain and high virulent strain was highly distinct.The pathogen of banana anthracnose latents on fruit, foliage, bract, blossom and stalk, stalk of plants with this disease is the most important carrier. The infection type of bannana fruit anthracnose is latent infection. The fruit of banana became carrier after 20 days of blossom, and the quantity of the pathogen increased gradually with the time prolong. After that the quantity of the pathogen of each growth phases related to rain fall. The the quantity of the pathogen of different parts was distinct.Infection process of banana fruit anthracnose pathogen was observed using free-hand section and paraffin section. Banana fruit were inoculated by conidia suspension, conidia germinated within 6hr, produced dilutus appressoria within 12hr,produced dematiaceous appressoria within 24hr. Appressoria were latent in intercellular cleft and were latent until banana fruit were harvested.The development process of conidia of Colletotrichum musae on fruit was not distinct from foliage and stalk.Two pairs of PCR primers were designed according to the two especial fragment (357bp and 206bp) of Colletotrichum musae. Banana tissue culture seedling genomic DNA, banana anthracnose pathogen genomic DNA, mango anthracnose pathogen genomic DNA, rubber anthracnose pathogen genomic DNA,watermelon anthracnose pathogen genomic DNA, banana crown rot pathogen genomic DNA, stylo anthracnose pathogen genomic DNA, watermelon Fusarium wilt pathogen genomic DNA were extracted using SDS method. Take the above genomic DNA as model, results from assays done with one pairs of primers showed that 357bp fragment was amplified from genomic DNA of C. musae while others not.So the pair of primers could be used to molecular detection of banana anthracnose.There was interference phenomena between banana template and its pathogen template. The target fragment couldn’t produce when banana template doubled its pathogen template. The target fragment produced when the above mixed templates were treated by enzyme hydrolysis(EcoR I ). The target fragment couldn’t produce when EcoR I was exchanged for Pst Hind.The target fragment produced when banana fruit were inoculated by high concentration conidia suspension, the target fragment couldn’t produce when banana fruit were inoculated by low concentration conidia suspension.更多还原