|本期目录/Table of Contents|

[1]张志霞 高 莉 刘倩倩 曹素娟 兰利琼 卿人韦**.适用于流式细胞术的三角褐指藻核DNA染色方法*[J].应用与环境生物学报,2018,24(06):1-8.[doi:10.19675/j.cnki.1006-687x.2018.01013]
 Zhang Zhi-xia,Gao Li,Liu Qian-qian,et al.The flow cytometric analysis of nucleus DNA stained with SYBR Green I in Phaeodactylum tricornutum*[J].Chinese Journal of Applied & Environmental Biology,2018,24(06):1-8.[doi:10.19675/j.cnki.1006-687x.2018.01013]
点击复制

适用于流式细胞术的三角褐指藻核DNA染色方法*()
分享到:

《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
24卷
期数:
2018年06期
页码:
1-8
栏目:
技术与方法
出版日期:
2018-12-25

文章信息/Info

Title:
The flow cytometric analysis of nucleus DNA stained with SYBR Green I in Phaeodactylum tricornutum*
文章编号:
201801013
作者:
张志霞 高 莉 刘倩倩 曹素娟 兰利琼 卿人韦**
四川大学生命科学学院 成都 610064
Author(s):
Zhang Zhi-xia Gao Li Liu Qian-qian Cao Su-juan Lan Li-qiong Qing Ren-wei**
College of Life Science, Sichuan University, Chengdu 610064, China
关键词:
三角褐指藻流式细胞术PISYBR-green I细胞核DNA染色
Keywords:
Phaeodactylum tricornutum flow cytometry propidium iodide SYBR-green I staining of nuclear DNA
DOI:
10.19675/j.cnki.1006-687x.2018.01013
摘要:
为制备可用于配置有488 nm激发器的流式细胞仪检测核DNA含量的三角褐指藻细胞,取对数生长期的三角褐指藻细胞,分别采用70%乙醇、1%戊二醛、1%甲醛溶液进行细胞固定,经RNase A消化后的样品用PI染色,用SYBR-green I对未经RNase A消化的细胞染色,在荧光显微镜下观察细胞染色情况,用流式细胞仪检测细胞核DNA含量。结果显示:在荧光显微镜下观察到,三种不同固定液固定的细胞均表现为SYBR-green I只对藻细胞核染色,PI对于藻的细胞核与细胞质均有着色。流式结果的DNA直方图显示,SYBR-green I染色的细胞的DNA直方图有明显的G1与G2/M峰,拟合度(RCS)在3左右,变异系数(%CV)在8.6左右。而PI染色细胞的DNA直方图表现为变异系数值偏大,拟合度较低,没有明显的G1与G2/M峰。因此,SYBR-green I是一种无需对细胞进行RNase A处理,可应用于配置有488 nm激光器流式细胞仪的核酸染料。(图5表2参25)
Abstract:
Objectives: The study aimed to investigate a suitable method to stain the nuclear of Phaeodactylum tricornutum applied to flow cytometry equipped with a 488-nm laser to detect the DNA content.Methods: Exponentially growing cells were fixed using 70% ethanol, 1% glutaraldehyde and 1% formaldehyde solution respectively.The samples were treated with RNase A and were then stained with propidium iodide. Those samples without RNase A treatment were stained with SYBR-green I. All these samples were observed by fluorescence microscope and the content of the nuclear DNA was quantitated by flow cytometry. Specific fluoresce spots indicate fluorescence is detected only in the nucleus of cells stained with SYBR-green I without RNase A treatment under fluorescence microscopy. Results: The result of flow cytometry indicates that the DNA histogram of cells stained with SYBR-green I demonstrates distinct G1 and G2 / M peaks, of which fit degree (RCS) is about 3, and the coefficient of variation (%CV) is around 8.6; however, the DNA histograms of cells stained with propidium iodide have a higher value of %CV, poor fit degree and no obvious G1 and G2/M peaks. Conclusion: The results indicate that SYBR Green I was adequate for nuclear DNA quantitation in Phaeodactylum tricornutum cells by flow cytometry equipped with a 488-nm laser without RNase A treatment.

相似文献/References:

[1]刘婷婷,陈南,孔维文,等.基于流式细胞术研究管网水中细菌的稳定性[J].应用与环境生物学报,2016,22(01):146.[doi:10.3724/SP.J.1145.2015.06011]
 LIU Tingting,CHEN Nan,KONG Weiwen,et al.Monitoring the bacterial stability in drinking water distribution systems by flow cytometry[J].Chinese Journal of Applied & Environmental Biology,2016,22(06):146.[doi:10.3724/SP.J.1145.2015.06011]

备注/Memo

备注/Memo:
收稿日期: 2018-01-10 接受日期: 2018-03-15
*国家自然科学基金(40976092)、四川省科技厅项目(2014JY0171)资助?
**通讯作者(E-mail: qingrw@scu.edu.cn)
点击摘要页题目后的“PDF”可下载阅读全文;本文为已录用的作者修定稿,尚未经编辑全面修改。
引用本文请注明出处本刊;发表刊期和页码将以正式出版时的安排为准,但DOI确定不变。
更新日期/Last Update: 2018-04-16