| 摘要: |
| 【目的】建立并优化小麦条锈菌的MFLP技术体系。【方法】以小麦条锈菌混合菌种为材料，通过对DNA提取、限制性内切酶2次酶切、PCR双重扩增等试验因素的选择优化，建立适用于小麦条锈菌的MFLP分析体系。【结果】改良的CTAB/SDS法有较好的破壁效果，所提取的DNA品质良好，完全符合MFLP的操作要求。优化确定的最佳MseⅠ酶切反应时间为4 h，预扩增最适退火温度为55 ℃，选择性扩增使用的模板为预扩增产物的100倍稀释物，其最适退火温度为58 ℃。【结论】 建立的MFLP技术体系可提供清晰、稳定的MFLP指纹图谱，为小麦条锈菌等专性寄生菌的群体遗传学与流行学研究提供了新的技术手段和方法。 |
| 关键词: 小麦条锈菌 MFLP 分子标记 |
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| 基金项目:河南省科技攻关计划项目（082102170024；092300410092）；国家“863”高新技术研究与发展计划项目（2006AA10Z429）；国家“十一五”科技支撑计划项目（2006BAD08A14） |
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| Establishment and optimization of the MFLP analysis of Puccinia striiformis f.sp.tritici |
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| Abstract: |
| 【Objective】 This study was intended to establish the MFLP reaction system special for Puccinia striiformis f. sp.tritici.【Method】Mixed uredospores of wheat stripe rust was used as the test material.Several main factors such as the genomic DNA preparation,restriction enzyme digestion and PCR amplification were studied and optimized.【Result】The improved CTAB/SDS method was so effective at cell wall breaking that the DNA product was qualified enough for later MFLP analysis.The optimal digestion time of MseⅠwas 4 h.The best annealing temperature of the preamplification was 55 ℃.As for the selective amplification,a 100-times dilution of the preamplification product was used as template and the preferable annealing temperature was 58 ℃.【Conclusion】The MFLP reaction system established in this study always gave the clear and stable MFLP fingerprint,which would provide a new research method for the obligated fungi,such as P.striiformis f.sp.tritici,in the field of population genetics and epidemiology. |
| Key words: Puccinia striiformis f.sp.tritici MFLP molecular marker |
