大肠杆菌共表达LeGGPS2、LePSY1和crtI基因合成番茄红素

Lycopene synthesis via tri-cistronic expression of LeGGPS2, LePSY1 and crtI in Escherichia coli

摘要:

利用大肠杆菌研究番茄红素的合成，不仅可以获得副产物少的高产菌株，而且可以探讨基因或基因簇的功能。文中将番茄LeGGPS2和LePSY1的cDNA序列，及欧文氏菌crtI的编码序列分别添加上核糖体结合位点后，以单独或组合的方式受控于T7启动子和终止子，在大肠杆菌菌株BL21 (DE3) 中进行表达和诱导番茄红素合成。结果显示，仅T7::crtI-LeGGPS2- LePSY1三价基因共表达时才能合成番茄红素，且将种子液以1∶50接种于含3％蔗糖的LB培养基 (pH 6.8) 中，于37 ℃摇8 h左右的对数

Abstract:

Studies on lycopene synthesis in Escherichia coli were not only able to gain the strains with high yield and less by-products, but also able to test functions of genes or gene clusters. In this article, the cDNA sequences of tomato LeGGPS2 and LePSY1 as well as the coding sequence of crtI from Erwinia uredovora, each of which was added a ribosome biding site, were controlled by T7 promoter and terminator alone or combined, and expressed in E. coli BL21 (DE3) to induce lycopene synthesis. The results show that only T7::crtI-LeGGPS2-LePSY1 expressed tri-cistronically could produce lycopene, and 2.124 mg/g dry cell weight of lycopene was obtained when fermented for 5 h at 30 ℃ after mixing 80 μmol/L IPTG at the later logarithmic phase while the seed broth of 1:50 (V/V) was inoculated into LB medium (pH 6.8) containing 3% sucrose and cultured for 8 h at 37 ℃. The results confirmed the function of the prokaryonized LeGGPS2 and LePSY1 and their synergy with crtI, and also laid a foundation to establish an independent lycopene synthetic pathway in the tomato plastid.