常绿阔叶林优势种福建青冈RAPD反应体系的建立

以福建青冈幼叶为材料,利用改进的CTAB法提取福建青冈幼叶的总DNA,探索适合于随机扩增多态DNA标记(RAPD)扩增的反应体系.结果表明,福建青冈合适的RAPD反应体系为:反应体积15μL,含10 ng模板DNA、1.5 mmo.lL-1MgC l2、0.3μmol.L-1引物、0.3 mmo.lL-1dNTP、1 UTaqDNA聚合酶.扩增程序为:94℃预变性3 m in,然后于94℃变性30 s,37℃复性30 s,72℃延伸90 s,循环41次,最后于72℃延伸7 m in.应用上述反应体系进行福建青冈的RAPD扩增反应,扩增产物进行12 g.L-1琼脂糖凝胶电泳,EB染色后用紫外凝胶成像系统分析,可获得满意的指纹图谱.

Establishment of RAPD reaction system of Cyclobalanopsis chungii, the dominant species of evergreen broad-leaf forest

An advanced CTAB method was used to extract the total DNA with the young leaves of Cyclobalanopsis chungii to establish an appropriate random amplified polymorphic DNA(RAPD) amplification system.The result showed that the optimal RAPD reaction system was with total volume of 15 μL,which consisted of 10 ng genomic DNA,1.5 mmol·L~(1) MgCl_2,0.3 μmol·L~(-1) dNTP,1 unit Taq polymerase.And the amplification procedure was as follows: preparatorily denaturing for 3 min at 94 ℃,denaturing for 30 sec at 94 ℃,annealing for 30 sec at 37 ℃,elongating for 90 sec at 72 ℃,41 cycles,and elongating for 7 min at 72 ℃.The C.chungii RAPD amplification reaction was used with the above reaction system,and the amplification products could get a pleasing finger map by 12 g·L~(-1) agarose gel electrophoresis and the ultraviolet gel imaging system analysis after EB dyeing.