山药染色体制片技术的优化

摘要

Chinese yam is a common vegetable food with the“medicine food homology”, while the research on the yam chromosome is far behind other vegetables. In this paper, the effects on chromosome squashing of different methods of gaining the roots, pretreatment solutions, pretreatment conditions and dissociation methods were studied to select the optimal squashing technique. The results showed that the best way to obtain root tips with a much higher number of metaphase cells was to cultivate yam tubers or bulbil in the lab. This method also obtained more metaphase cells which chromosomes were separated well. About pretreatment conditions,8-hydroxyquinoline or the mixture of 8- hydroxyquinoline and colchicine as the pretreatment liquid was suitable for chromosome karyotype analysis. The best pretreatment method was to treat the root tips in 2 mmol/L 8-hydroxyquinoline liquid for 2 h at 4℃. The enzymatic and acid dissociation method was better than the acid dissociation method. The former method showed the ideal dissociation effect with the clear chromosomes, and the chromosomes of 96.3%mitotic metaphase cells were in one horizontal plane.%山药(Dioscorea opposite Thunb.)为药食兼用的常见蔬菜，而其染色体的研究相对滞后。以根尖为试验材料，分别考察不同的取材方式、预处理试剂、预处理方法以及解离方式对山药染色体制片效果的影响，筛选优化制片的关键技术，以期为今后细胞学研究打下基础。结果表明：室内基质培养块茎(或零余子)的方式获取的根尖中中期分裂相个数最多，且染色体分散良好的中期分裂相占的比例最高；适合于山药核型分析的预处理试剂为8-羟基喹啉同秋水仙素的混合液、8-羟基喹啉溶液，其中8-羟基喹啉溶液预处理后染色体缢痕更清晰，为最好的预处理试剂；预处理方法以低温4℃下2 mmol/L的8-羟基喹啉预处理2 h最好；采用酸解加酶解的方式解离效果明显好于单用酸解的方式，并且得到的制片中染色体形态清晰，96.3%的中期细胞的染色体分布于同一个水平面上。