连续光照转暗培养联合药物处理实现衣藻细胞的高水平同步化

刘维仲, 王洁茹, 胡勇

doi:10.3724/SP.J.1259.2015.00363

植物学报 >

2015 , Vol. 50 >Issue 3: 363 - 371

DOI: https://doi.org/10.3724/SP.J.1259.2015.00363

技术方法

连续光照转暗培养联合药物处理实现衣藻细胞的高水平同步化

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1山西师范大学生命科学学院, 临汾 041004；
2.首都师范大学生命科学学院, 北京 100048

收稿日期: 2014-12-25

修回日期: 2015-03-19

网络出版日期: 2015-04-08

基金资助

国家自然科学基金(No.31171167)和山西省自然科学基金(No.2009011044-1)

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Highly Synchronized Chlamydomonas reinhardtii Cells by Dark-shifting a Continuous-light Culture Combined with Drugs

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1College of Life Sciences, Shanxi Normal University, Linfen 041004, China;
2.College of Life Sciences, Capital Normal University, Beijing 100048, China

Received date: 2014-12-25

Revised date: 2015-03-19

Online published: 2015-04-08

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摘要

单细胞衣藻(Chlamydomonas)是光合作用和植物细胞周期等生物学过程研究的一个重要模式系统, 同步化培养是进行相关研究的必要手段。该研究探索了连续光照转暗培养联合细胞周期阻断剂实现莱茵衣藻(Chlamydomonas reinhardtii)细胞高水平同步化的新方法, 并利用流式细胞术对同步化程度进行了精确的分析。结果表明, 连续光照转暗培养或联合S期阻断剂可以使衣藻细胞同步化到G1期或G1/S期边界; 连续光照转暗培养联合M期阻断剂或者在“加入-释放”S期阻断剂后再加入M期阻断剂可以使衣藻细胞同步化到M期, 同步化水平可达80%。具体的同步化培养步骤要根据研究对象(特别是某些衣藻突变株系)的特性和研究目的确定。

本文引用格式

刘维仲, 王洁茹, 胡勇 . 连续光照转暗培养联合药物处理实现衣藻细胞的高水平同步化[J]. 植物学报, 2015 , 50(3) : 363 -371 . DOI: 10.3724/SP.J.1259.2015.00363

Abstract

Chlamydomonas reinhardtii, a unicellular eukaryotic green alga, has been a powerful model system for understanding several fundamental biological processes such as photosynthesis and plant cell-cycle regulation. An artificial cell-cycle synchronization system is necessary for these studies. Here, we investigated a new method for synchronizing Chlamydomonas cell-wall-less cells, with which a high level of synchrony can be achieved. The degree of synchrony was analysed by flow cytometry. The cells are grown asynchronously in continuous light, then grown for 24 h in the dark (dark-shifting a continuous-light culture). At the end of the dark period, most cells (>80%) are arrested in the G1 phase. If necessary, an S-phase blocking agent can be used to treat the dark-shifted cells. When the dark-shifted cells are treated with M-phase blocking agent alone or with the S-phase and M-phase blocking agent sequentially, a cell population with more than 80% of cells at the M phase can be achieved. A specific synchronous procedure needs to be selected according to the properties of a certain Chlamydomonas strain (especially certain mutants) and the requirement of research.

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