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构建染色体特异文库是简化小麦基因组测序的有效途径,而通过同步化处理提高根尖细胞有丝分裂指数,使根尖细胞中富集高比例的中期染色体是实现染色体分选成功的前提。采用双阻断法用50μmolL?1羟基脲和100μmolL?1氟乐灵对普通小麦根尖细胞进行同步化处理,然后用流式核型分析法观测各周期细胞在不同时间点上的比例变化。结果显示,最佳方案为羟基脲处理10h,恢复7h,氟乐灵处理4h。以此方案处理中国春小麦根尖可获得最高的有丝分裂指数,达70.1%。附加冰水处理(0℃)有助于减少染色体碎片和增加流式核型的分辨率,但对富集更多的中期染色体没有效果。
The construction of chromosome-specific library is an effective way to simplify the sequencing of wheat genome. To improve the mitotic index of root tip cells through synchronized processing, enriching the root tip cells with high proportion of metaphase chromosomes is the premise of chromosome success. The double-stranded method was used to synchronize the root tip cells of common wheat with 50μmol L -1 hydroxyurea and 100μmol L? Trifluralin, then the flow cytometry was used to observe the changes of the proportion of cells in different periods. The results showed that the best solution for hydroxyurea treatment 10h, recovery 7h, trifluralin treatment 4h. With this scheme, the highest mitotic index of Chinese spring wheat root tips was obtained, reaching 70.1%. Additional ice water treatment (0 ° C) helps to reduce chromosomal fragmentation and increase the resolution of flow cytometry, but has no effect on enriching for more metaphase chromosomes.