| 谢家楠，郭建军,金道超*.白背飞虱ISSR-PCR反应体系构建的正交设计优化[J].植物保护,2014,40(2):109-112. |
| 白背飞虱ISSR-PCR反应体系构建的正交设计优化 |
| Establishment and optimization of ISSR PCR reaction system with orthogonal design for Sogatella furcifera |
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| DOI： |
| 中文关键词: 白背飞虱 正交设计 ISSR |
| 英文关键词:Sogatella furcifera orthogonal design ISSR |
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| 摘要点击次数: 1049 |
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| 中文摘要: |
| 白背飞虱是水稻重要害虫之一。研究采用L16(45)正交设计优化试验，构建了白背飞虱ISSR最优反应体系，并对引物退火温度和反应循环次数进行了优化。结果表明：白背飞虱ISSR PCR最优反应体系为2 μL 10×PCR Buffer（10 mmol/L Tris HCl;50 mmol/L KCl）,1.5 mmol/L Mg2+,0.25 mmol/L dNTPs，0.9 μmol/L引物，1U TaqPolymerase和50 ng 模板DNA，ddH2O补充至20 μL。本研究采用的10条引物的最佳退火温度在51.0~54.0 ℃间，以引物相应最优退火温度和40次反应循环可获得较好的白背飞虱ISSR-PCR结果。 |
| 英文摘要: |
| Sogatella furcifera is one of the most important rice pests. To establish and optimize ISSR PCR reaction system for S.furcifera, the factors influencing ISSR system were studied with L16(45) orthogonal design, and the optimal annealing temperature and cycle times were proposed. The results showed that the optimal conditions for ISSR PCR system were 2 μL 10×PCR buffer (10 mmol/L Tris HCl, 50 mmol/L KCl), 1.5 mmol/L Mg2+, 0.25 mmol/L dNTPs mixture, 0.9 μmol/L of each primer, 1U TaqDNA polymerase (TaKaRa) and 50 ng template DNA in 20 μL reaction volume. The optimal annealing temperatures were 51.0-54.0 ℃ for the 10 primers used in the present study. The clear and reproducible DNA bands were obtained by the optimal annealing temperature and 40 reaction cycles. |
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