单粒蔬菜种子DNA快速提取方法的建立OA北大核心CSTPCD
Establishment of Method for Quick Extraction of DNA from Single Vegetable Seed
利用磁珠法对甘蓝、白菜、芥菜、洋葱、黄瓜、番茄、辣椒等7种形状、大小不同的蔬菜单粒种子和幼苗进行DNA提取,采用内参基因Actin进行实时荧光定量PCR评价DNA质量,利用KASP(kompetitive allele specific PCR)标记检验DNA质量是否达到KASP检测要求.结果显示,单粒种子磁珠法提取的各蔬菜DNA平均Ct值均处于19～24范围内.KASP检测进一步证实种子提取DNA的质量达到KASP标记分型的要求.综上所述,本研究建立了一种简单、经济、实用性强的单粒种子DNA提取方法,该方法对不同大小的蔬菜种子具有较好的稳定性,且大幅缩短了获得基因型数据的时间.
DNA was extracted from single seed as well as seedlings of seven vegetables,including of cabbage,Chinese cabbage,mustard,onion,cucumber,tomato and pepper using magnetic bead method.The seeds of these seven vegetables varied in size and shape.The DNA quality was evaluated by real-time fluorescent PCR reaction using reference gene Actin.In order to check whether the DNA quality was good enough for KASP(kompetitive allele specific PCR)assay,we conducted KASP assay for each vegetable.The results showed that the average Ct value of DNA extracted from single vegetable seed by magnetic beads was in the range of 19-24.KASP assays further confirmed their quality was good enough for KASP markers assay.In conclusion,this study established a simple,economical and practical method for DNA extraction of single seeds,which has good stability for vegetable seeds of different sizes and greatly reduces the time for obtaining genotype data.
杨帆;武剑;常立春;张涛;朱海峰;梁建丽;高杰;王晓武
新疆农业大学园艺学院,新疆特色园艺作物种质资源与高效生产实验室,新疆乌鲁木齐 830052||中国农业科学院蔬菜花卉研究所,蔬菜生物育种全国重点试验室,北京 100081中国农业科学院蔬菜花卉研究所,蔬菜生物育种全国重点试验室,北京 100081中国农业科学院蔬菜花卉研究所,蔬菜生物育种全国重点试验室,北京 100081||山西农业大学,山西晋中 030801新疆农业大学园艺学院,新疆特色园艺作物种质资源与高效生产实验室,新疆乌鲁木齐 830052
蔬菜种子磁珠法DNA提取KASP分子标记实时荧光定量PCR
vegetable seedmagnetic bead DNA extractionKASP markerreal-time fluorescence quantitative PCR
《中国蔬菜》 2024 (007)
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财政部和农业农村部:国家现代农业产业技术体系项目(CARS-10-P19),新疆维吾尔自治区园艺学重点学科建设项目(2016-10758-37),中国农业科学院基本科研业务费专项(Y2023PT16),国家重点研发计划项目(2022YFF1003003)
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