Purification of the recombinant green fluorescent protein from tobacco plants using alcohol/salt aqueous two-phase system and hydrophobic interaction chromatography

BMC Biotechnology

Sheng Wang¹  Xiangzhen Ding¹  Jie Dong¹ 

[1] Key Laboratory of Ministry of Education for Protection and Utilization of Special Biological Resources in the Western China;

关键词: Green fluorescent protein;    Plant virus;    Transient gene expression;    Aqueous two-phase system;    Hydrophobic interaction chromatography;   

DOI  :  10.1186/s12896-019-0590-y

来源: DOAJ

【 摘 要 】

Abstract Background The green fluorescent protein (GFP) has been regarded as a valuable tool and widely applied as a biomarker in medical applications and diagnostics. A cost-efficient upstream expression system and an inexpensive downstream purification process will meet the demands of the GFP protein with high-purity. Results The recombinant GFP was transiently expressed in an active form in agoinoculated Nicotiana benthamiana leaves by using Tobacco mosaic virus (TMV) RNA-based overexpression vector (TRBO). The yield of recombinant GFP was up to ~ 60% of total soluble proteins (TSP). Purification of recombinant GFP from the clarified lysate of N. benthaniana leaves was achieved by using an alcohol/salt aqueous two-phase system (ATPS) and following with a further hydrophobic interaction chromatography (HIC). The purification process takes only ~ 4 h and can recover 34.1% of the protein. The purity of purified GFP was more than 95% and there were no changes in its spectroscopic characteristics. Conclusions The strategy described here combines the advantages of both the economy and efficiency of plant virus-based expression platform and the simplicity and rapidity of environmentally friendly alcohol/salt ATPS. It has a considerable potential for the development of a cost-efficient alternative for production of recombinant GFP.

【 授权许可】

Unknown