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中国农学通报

中国农学通报 ›› 2009, Vol. 25 ›› Issue (21): 244-247.

• 林学 园艺 园林 • 上一篇    下一篇

湖南宽皮柑橘EST-SSR反应体系研究

曾柏全1,2,邓子牛1,熊兴耀1,杨迎花1   

  • 收稿日期:2009-06-09 修回日期:2009-07-20 出版日期:2009-11-05 发布日期:2009-11-05
  • 基金资助:

    新型分子标记研发及在湖南宽皮柑橘中应用

Study on the EST-SSR Marker Reaction System of mandarins(Citrus Reticulata Blanco) in Hunan

  • Received:2009-06-09 Revised:2009-07-20 Online:2009-11-05 Published:2009-11-05

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摘要：

首次以EST-SSR 技术应用于湖南宽皮柑橘的研究中，进行了引物的设计与筛选，建立了湖南宽皮柑橘的EST-SSR 的反应体系。共筛选了20对引物，其中12对引物扩增多态性较高，研究确定在25 uL的反应体系中, Mg2+浓度在2.5 mmol?L-1, dNTP 0.2 mmol?L-1,上下引物各为0.2umol?L-1, Taq酶浓度为1.0 U效果好 , 最佳PCR程序为94℃预变性１ min，然后94 ℃，1 min；52 ℃，1min；73 ℃，1min，45个循环，最后73 ℃延伸3 min,利用优化后的反应体系成功构建湖南宽皮柑橘EST-SSR的指纹图谱.

关键词: 水分胁迫, 水分胁迫, 橡胶树

Abstract:

It was the first time to study mandarins(Citrus Reticulata Blanco ) with EST-SSR technology , clean and stable bands had been detected by twelve pairs of EST-SSR primer which had been selected out from twenty pairs .Each 25ul PCR reaction mixture consisted of genemic DNA , 2.5 mmol?L-1 of MgCl2, 0.2 mmol?L-1 of dNTP, 0.2u mol?L-1 of enemprimer and 1.0 unit of Taq polymerase .The results showed that the most suitable protocol was initially denaturing at 94℃ for 1 min; then 94℃,60℃second; 52℃, 60 second;and 73℃，1min for 45 cycles.，and finally extended at 73 ℃ for 3 min . It was established the genetic linking maps of mandarins in Hunan with optimizal reaction system.