多倍体西瓜染色体制片技术改良及其核型分析  

Optimization of chromosome preparation and karyotype analysis of polyploid watermelons

作　　者：李桂芬[1] 何毅[1] 覃斯华[1] 黄金艳[1] 柳唐镜[1] 洪日新[1] 李天艳[1] 樊学军[1] 韦正光[1] 李文信[1] 

机构地区：[1]广西农业科学院园艺研究所,南宁530007

出　　处：《南方农业学报》2017年第4期663-668,共6页Journal of Southern Agriculture

基　　金：国家西甜瓜产业技术体系建设项目(CARS-26);广西自然科学基金项目(2013GXNSFBA019099);广西农业科学院科技发展基金项目(桂农科2013JZ02;桂农科2015JZ013)

摘　　要：【目的】改良多倍体西瓜染色体标本制片技术,比较多倍体西瓜常规核型分析与荧光核型分析结果,为西瓜细胞遗传学研究及辅助构建西瓜物理遗传图谱提供参考依据。【方法】以多倍体西瓜为材料,以传统染色体制片方法去壁—低渗火焰干燥法为基础,去掉其前低渗和后低渗两个步骤,合并其再固定与火焰涂片两个步骤,用吉姆萨染色剂和荧光染料DAPI分别对多倍体西瓜染色体进行常规染色和荧光染色,并进行核型分析。【结果】使用改良型酶解去壁火焰干燥法进行多倍体西瓜染色体制片,比传统去壁—低渗火焰干燥法简化了用KCl溶液前低渗、后低渗及用卡诺固定液再固定3个步骤,可缩短制片时间1.0~2.0 h,提高制片效率,获得的染色体样本数目齐全、分散良好、形态清晰。用DAPI染色比用吉姆萨染色更容易检测到细胞中期分裂相,每张玻片可缩短检测时间0.5~1.5 h;染色体的着丝点、长臂及短臂形态更清晰。【结论】采用改良染色体制片方法可提高多倍体西瓜染色体的制片效率,对该制片进行荧光核型分析比常规核型分析更准确,效率更高。[Objective]Technique of chromosome preparation for polyploid watermelon was improved,conventional karyotype and fluorescence karyotype for polyploid watermelons were compared in order to provide reference for research on genetics of watermelon and establishment of physical genetic map of watermelon. [ Method ] Polyploid watermelons were used as materials in the study. Based on traditional chromosome preparation technique, namely wall degradation-hypotonic flaming-drying method, pre-hypotonie and afler-hypotonic were deleted, and re-fixation and flame smear were combined into one procedure. Giemsa stain and DAPI fluorescence stain were used to stain, and carried out conventional karyotype analysis and fluorescence karyotype analysis respectively. [ Result]The optimized enzymolysis flaming-drying method was used to prepare chromosome of polyploid watermelons. Compared with the traditional technique, the optimized one simplified three procedures which were pre-hypotonic and after-hypotonic of KCL solution and re-fixation by Carnoy fixative. Thus the time was 1.0-2.0 h shorter. Production efficiency was improved, and samples with complete amount, good dispersion and clear configuration were obtained. Slides stained with DAPI could shorten the detection time for 0.5-1.5 h each slide, and were more easily to be detected metaphases with more clear centromere,long arm and short arm of chromosome than stained with Giemsa. [ Conclusion ] The modified method can improve the efficiency of chromosome preparation of polyploid watermelon. Fluorescent karyotype analysis is more accurate and efficient than conventional karyotype analysis.

关 键 词：多倍体西瓜 染色体制片 荧光染色 核型分析 

分 类 号：S651[农业科学—果树学]