蜡梅AFLP分子标记技术体系的建立

The Establishment of AFLP Molecule Labeling Technique System of the Wintersweet Cultivars

利用简易CTAB法、改良的CTAB法和SDS法提取蜡梅[Chimonanthus praecox(L.)Link]成熟叶和嫩叶的基因组,并进行了检测比较。结果显示,改良的CTAB法更适合蜡梅基因组DNA的提取,蜡梅叶片的年龄并不影响蜡梅基因组DNA的提取;同时利用AFLP分子标记技术,采用MseI-EcoR I酶切组合,从168对引物中筛选出10对带型分布均匀、多态性高且分辨能力强的引物,分别为:M23E46、M24E46、M25E46、M23E47、M24E47、M41E47、M41E94、M64E94、M64E66和M24E75,并确定了适用于蜡梅AFLP反应的最佳酶切连接、预扩和选扩体系,从而为今后利用AFLP分子标记技术研究蜡梅的品种分类和野生居群的遗传多样性分析打下坚实的基础。

The methods of CTAB, improved CTAB and SDS were used for genomic DNA isolation of climax leaves and young leaves of Wintersweet cultivars. The results indicated that the methods of CTAB was useful in Wintersweet cultivars genomic DNA isolation, and the leaf age doesn＇ t affect genomic DNA isolation of Wintersweet cultivars. By the amplified fragment length polymorphisms （AFLP） technique, using MseI-EcoR I enzyme-cut combination, ten pairs of primers （ i.e. M23E46, M24E46, M25E46, M23E47, M24E47, M41E47, M41E94, M64E94, M64E66 and M24E75） with high polymorphism and powerful distinctiveness were selected from 168 pairs of primer, and the best pre-expanding and selective expanding system were determined. Our results tentatively established the basis for the research of the cuhivar classification and gentic diversity analysis.