基于流式细胞仪鉴定马铃薯倍性的高通量样品制备方法
High-throughput Sample Preparation Method for the Identification of Potato Ploidy Using Flow Cytometry
袁兰 1黄娅楠 2张贝妮 1熊雨萌 3王洪洋3
作者信息
- 1. 云南师范大学云南省马铃薯生物学重点实验室,昆明 650500;云南师范大学生命科学学院,昆明 650500
- 2. 云南师范大学生命科学学院,昆明 650500
- 3. 云南师范大学云南省马铃薯生物学重点实验室,昆明 650500
- 折叠
摘要
[目的]染色体倍性鉴定是马铃薯种质资源评价的重要内容.建立马铃薯染色体倍性的高通量样品制备方法,可为后续大规模开展马铃薯倍性鉴定工作奠定基础.[方法]以 30 份马铃薯孤雌生殖诱导后代作为实验材料,比较使用低温钢珠打样法、液氮研磨法和刀片切碎法制备细胞核悬液的效果,并对已知二倍体马铃薯IVP101 和四倍体马铃薯HTJ349-3 进行倍性鉴定.[结果]低温钢珠打样法制备的细胞核悬液,荧光信号明显,细胞裂解充分,杂质少.同时,由于低温钢珠打法简单易操作,大大缩短了样品制备时间,其鉴定效率比起液氮研磨法提高 45-60 倍,比刀片切碎法提高 105-180 倍.利用该方法可以准确检测出马铃薯IVP101 和HTJ349-3 的染色体倍性.[结论]基于流式细胞仪的低温钢珠打样法,其检测结果同液氮研磨法、刀片切碎法制样方法一样准确,可实现高通量鉴定马铃薯染色体倍性.
Abstract
[Objective]Chromosome ploidy identification is an important part of the evaluation of potato germplasm resources.The establishment of a high-throughput sample preparation method for the identification of potato chromosome ploidy based on flow cytometry would lay a foundation for the subsequent large-scale identification of potato ploidy.[Method]The 30 parthenogenetic offspring of potato were used to compare the effects of low-temperature steel-bullet beating,liquid nitrogen grinding and blade shredding to prepare nuclear suspensions.The ploidy of diploid potato IVP101 and tetraploid potato HTJ349-3 was identified.[Result]The suspension of cell nuclei prepared by the low-temperature steel-bullet beating method had obvious fluorescence signal,sufficient cell lysis and few impurities.Due to the simple and easy operation of the low-temperature steel-bullet beating method,the sample preparation time was greatly shortened,and the identification efficiency was 45-60 times higher than that of the liquid nitrogen grinding method,and 105-180 times higher than that of the blade shredding method.The chromosome ploidy of potato IVP101 and HTJ349-3 was accurately detected by this low-temperature steel-bullet beating method.[Conclusion]The results by the low-temperature steel-bullet beating method are as accurate as that by the liquid nitrogen grinding method and the blade cutting method,which can achieve high-throughput identification of potato chromosome ploidy.
关键词
流式细胞术/马铃薯/倍性鉴定/低温钢珠打样法Key words
flow cytometry/potato/ploidy identification/low-temperature steel-bullet beating method引用本文复制引用
基金项目
国家自然科学基金项目(32360523)
国家自然科学基金项目(32060499)
云南省基础研究计划项目面上项目(202201AT070034)
云南省兴滇英才支持计划青年人才项目(XDYC-QNRC-2022-0218)
出版年
2024
国家科技期刊平台
NSTL
万方数据