Quantitation of Cell-Cell Fusion Using the Multicolor FCM Assay I

The purpose of the experiment is to quantify cell-cell fusion by flow cytometric analysis of a wt X wt and a prm1∆ X prm1∆ yeast mating. MATa and MATalpha strains are distinguished by staining each strain with ConcanavalinA- Alexa Fluor 647 (ConA-647) and with ConcanavalinA- Tetramethylrhodamine (ConA-Tet), respectively. Mating pairs are revealed as two-colored entities. Cytoplasmic mixing is measured with a GFP bi-molecular fluorescence complementation assay. Cell fusion efficiency is calculated as the percentage of fused mating pairs over the total number of pairs.
Conclusion:
Cell fusion efficiencies of a wt X wt mating and a prm1∆ X prm1∆ mating were quantified. Each mating sample was analyzed by triplicate, unstained and compensation controls are included in the experiment.

本实验旨在通过流式细胞分析（flow cytometric analysis），定量检测野生型（wild type，wt）×野生型与prm1∆×prm1∆酵母交配体系的细胞融合效率。其中，MATa与MATα型菌株分别通过刀豆蛋白A-亚历克斯荧光647（ConcanavalinA- Alexa Fluor 647，ConA-647）与刀豆蛋白A-四甲基罗丹明（ConcanavalinA- Tetramethylrhodamine，ConA-Tet）染色进行区分。交配伴侣呈现为双色信号实体。细胞质混合情况通过绿色荧光蛋白（green fluorescent protein，GFP）双分子荧光互补实验进行检测。细胞融合效率以融合交配伴侣数占总交配伴侣数的百分比进行计算。
结论：
本实验定量检测了野生型×野生型与prm1∆×prm1∆酵母交配体系的细胞融合效率。所有交配样本均设置三次重复实验，实验同时包含未染色对照与补偿对照。