| 油松SSR-PCR引物筛选及反应体系的建立 |
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| 引用本文： | 张冬梅,杨娅,沈熙环,茹广欣.油松SSR-PCR引物筛选及反应体系的建立[J].北京林业大学学报,2007,29(2):13-17. |
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| 作者姓名： | 张冬梅 杨娅 沈熙环 茹广欣 |
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| 作者单位： | 上海市园林科学研究所;河南职业技术学院;北京林业大学生物科学与技术学院;河南农业大学林学园艺学院 |
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| 摘 要： | 油松是我国特有的重要乡土针叶树种，开发合适的油松PCR-SSR引物并建立优化的反应体系，是开展油松天然群体和种子园人工群体遗传研究的基础.该研究以油松总DNA为材料，分析了Taq聚合酶、样本浓度、Mg2+浓度、dNTP浓度以及引物浓度对PCR SSR扩增结果的影响，筛选出扩增条带清晰、多态性丰富的SSR引物12对，建立了稳定的、可重复的油松PCR-SSR最佳反应体系及PCR扩增参数.研究结果表明:在15 μL SSR -PCR反应体系中，样本最适宜浓度为30 ng，Mg2+的最适浓度为0.25 mmol/L，dNTP最适浓度为0.2 mmol/L，单引物的最适浓度均为250 nmol/L；Taq聚合酶在15 μL反应体系中宜加入0.375 U.统计利用所选12对引物，对辽宁兴城油松种子园内49个建园无性系进行了SSR-PCR反应，通过6%的变性聚丙稀酰胺凝胶电泳检测，每对引物扩增产物在100～250 bp之间的等位谱带数最大为10，最小为5，不同无性系间DNA谱带多态性丰富.油松SSR-PCR引物筛选及反应体系的建立，为今后利用SSR标记技术开展油松种子园的父本分析及选择性受精研究提供了一个标准化程序和强有力的工具.
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| 关 键 词： | 油松 SSR标记 引物 反应体系 优化 |
| 文章编号： | 1000-1522（2007）02-0013-05 |
| 收稿时间： | 1900-01-01 |
| 修稿时间： | 2006-02-13 |
Selection of primers and establishment of SSR-PCR reaction system on Pinus tabulaeformis Carr |
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| ZHANG Dong-mei,YANG Ya,SHEN Xi-huan,RU Guang-xin.Selection of primers and establishment of SSR-PCR reaction system on Pinus tabulaeformis Carr[J].Journal of Beijing Forestry University,2007,29(2):13-17. |
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| Authors: | ZHANG Dong-mei YANG Ya SHEN Xi-huan RU Guang-xin |
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| Institution: | 1.Shanghai Landscape Gardening Research Institute, 200232, P. R. China; 2 Henan Vocational Technology Institute, Zhengzhou, 450046, P. R. China ; 3 .College of Biological Sciences and Bioteehnology, Beijing Forestry University, 100083, P. R. China; 4. College of Forestry and Horticulture, Henan Agricultural University, Zhengzhou, 450002, P. R. China. |
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| Abstract: | Taking Pinus tabulaeformis Carr. as the material, the effects of primer pairs, whole DNA concentration, Mg2+and Taq polymerase concentration and dNTP concentration on SSR reaction system were studied. Twelve primer pairs with abundant polymorphism bands were selected and the SSR PCR reaction system on P. tabulaeformis was established too. The results showed that in 15 μL PCR reaction: 30 ng DNA, 0.25 mmol/L Mg2+, 0.2 mmol/L dNTP, 250 nmol/L Primer(F), 250 nmol/L Primer(R), 0375 U Taq polymerase were the best. Detected by 6% polyacryl amide gel, polymorphism among 49 clones of P. tabulaeformis in the Seed Orchard of Xingcheng, Liaoning Province was abundant, the length of amplification products was 100-250 bp, the number of alleles per locus varied from five to ten. The work provides a powerful tool for paternity analysis in P. tabulaeformisseed orchards. |
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| Keywords: | Pinus tabulaeformis Carr SSR marker primers reaction system optimization |
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