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油茶SSR-PCR反应体系建立与优化
刘冰,金龙,曹翠萍,王勤,周明善,吴山忠,束庆龙,张良富
0
(安徽农业大学林学与园林学院,合肥 230036;安徽省歙县特种经济林场,歙县 245900)
摘要:
油茶是我国特有的木本食用油料树种,本研究采用正交试验和单因素试验,建立并优化了油茶SSR-PCR 反应体系。结果表明,油茶最适SSR-PCR 反应体系为模板 DNA (5~10 ng·μL-1) 1.0 μL,Mg2+ (25 mmol·L-1) 0.7 μL,dNTP (10 mmol·L-1) 0.15 μL,正反向引物均为 (10 μmol·L-1) 0.2 μL,Taq DNA 聚合酶 (5 U·μL-1) 0.1 μL,10×PCR 缓冲液1.0 μL,HPLC水6.65 μL,总体积10.0 μL;同时证明了微卫星位点在山茶属 (Camellia L.) 植物的种间通用性。研究结果将为油茶遗传多样性研究、品种鉴定、亲缘关系分析以及重要农艺性状的QTL研究提供重要的理论依据和技术支持。
关键词:  油茶  微卫星标记  PCR反应体系  优化
DOI:CNKI:34-1162/S.20111025.1027.015
基金项目:安徽省人才开发专项资金(2008Z023), 安徽农业大学引进与稳定人才科研资助(yj2008-3), 安徽省自然科学基金(090411025)共同资助。
Establishment and optimization of SSR-PCR reaction system for oil tea (Camellia oleifera Abel.)
LIU Bing,JIN Long,CAO Cui-ping,WANG Qin,ZHOU Ming-shan,WU Shan-zhong,SHU Qing-long,ZHANG Liang-fu
(School of Forestry & Landscape Architecture,?Anhui Agricultural University,?Hefei 230036;Special Economic Forestry Centre of Shexian County of Anhui Province, Shexian 245900)
Abstract:
Camellia oleifera Abel. is an edible oil tree species native to China. SSR-PCR reaction system for C. oleifera was established and optimized through orthogonal test and single factor experiment. The results revealed that the optimal SSR-PCR reaction system was 10.0 μL volume containing DNA (5-10 ng·μL-1) 1.0 μL, Mg2+ (25 mmol·L-1) 0.7 μL, dNTP (10 mmol·L-1) 0.15 μL, F- and R-primer (10 μmol·L-1) 0.2 μL, respectively, Taq DNA (5 U·μL-1) 0.1 μL, 10×PCR buffer 1.0 μL, HPLC H2O 6.65 μL. The cross-species transferability of SSR loci within genus Camellia was validated. Information of the study may offer theoretical base and technical support to assessment of genetic diversity, identification of variety, phylogenetic study and QTL analysis of C. oleifera in the future.
Key words:  Camellia oleifera Abel.  SSR  PCR reaction system  optimization

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