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《Molecular Plant Breeding》 2008-01
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Establishment and Optimization of SRAP and ISSR Marker System in Citrus

Wu Xin 1,2,3 Lei Tiangang 2,3 He Yongrui 2,3 Liu Xiaofeng 2,3 Xu Lanzhen 2,3 Peng Aihong 2,3 Chen Shanchun 2,3 1 College of Horticulture & Landscape Designing, Southwest University, Chongqing, 400716; 2 Citrus Research Institute, Southwest University, Chongqing, 400712; 3 National Citrus Cultivar Improvement Center of China, Chongqing, 400712  
Here we have established SRAP-PCR and ISSR-PCR system in Citrus by optimization of PCR reaction program, reaction system (template DNA, reaction volume, Mg2+, dNTP, Taq DNA polymerase and primer) and electrophoresis detection experiments. Based on these reaction systems, a mass of primers have been screened. In the end, we got optimum SRAP and ISSR marker systems in Citrus. SRAP-PCR system: template DNA 25 ng, Tris-HCl 10 mmol/L, KCl 50 mmol/L, Mg2+ 1.2 mmol/L, dNTP 120 μmol/L, Taq DNA polymerase 1.5 U, primer 0.4 μmol/L in 25 μL system, reaction program: initial denaturation step for 5 min at 94℃, followed by 35 cycles at 94℃ for 30 s, 47℃ for 1 min and 72℃ for 1 min, followed by a final extension step at 72℃ for 10 min; ISSR-PCR system: template DNA 25 ng, Tris-HCl 10 mmol/L, KCl 50 mmol/L, Mg2+ 2.0 mmol/L, dNTP 200 μmol/L, Taq DNA polymerase 1 U, primer 0.8 μmol/L in 25 μL system. 24 pairs of SRAP primers and 13 ISSR primers, with high stability and polymorphism, has been obtained.
【Fund】: 科技部科研院所社会公益研究专项(2004DIB45147);; 重庆市科技攻关重点项目(CSTC 2006AB1009);; 国家863项目(2006AA100108-4-12-4)资助
【CateGory Index】: S666
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