红蓝荧光细胞的构建

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Abstract Objective: To construct the eukaryotic expression plasmid of BFP-H2B and express it in the B16 mouse melanoma cells which can already express red fluorescence protein (RFP) in the cytoplasm, and make the cells’ nucleus show blue light while the plasmid show red light when activated by suitable light wave. Methods: The blue fluorescence protein (BFP) was acquired with PCR from a constructed PAAV3, and H2B gene was obtained with RT-PCR from cultured 293 cell’s total RNA. The sequence encoding H2B and the sequence encoding BFP were inserted into pRetroX to construct the eukaryotic expression vector pRetroX-BFP-H2B. After the constructed plasmid was confirmed by enzymatic digestion and PCR digestion, the recombination plasmid was transfected into B16 mouse melanoma cells which could already express RFP in the cytoplasm by the lipofectamine transfection technology for observation of the fusion protein’s expression. Results: The transfection of double fluorescent cells could present a double fluorescent effect, and stably expresssd batches after successfully constructed the eukaryotic expression plasmid of BFP-H2B. It provided a new reference for the construction of double fluorescent cells and the selection of the fluorescent protein species. Conclusion: In this study we establish an available survey system for the study of the two color fluorescence cells and the selection of fluorescence proteins used.

Key words： gene H2B blue fluorescence protein melanoma cells eukaryotic expression plasmid

Received: 15 December 2014

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	WANG Zhen
	SHI Xinyan
	CHEN Yu.

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https://xb.wmu.edu.cn/EN/ OR https://xb.wmu.edu.cn/EN/Y2015/V45/I12/894

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