本文選題：草莓　切入點：FaMYB　出處：《基因組學(xué)與應(yīng)用生物學(xué)》2017年01期

【摘要】：利用中心組合設(shè)計(CCD)探究p ET-32a(+)-Fa MYB5融合蛋白原核表達的最優(yōu)條件。以不同的誘導(dǎo)溫度、誘導(dǎo)時間、菌液初始濃度和異丙基-β-D-硫代半乳糖苷(IPTG)終濃度為4個考察因素,以可溶性蛋白含量為考察指標,在大腸桿菌中表達p ET-32a(+)-Fa MYB5融合蛋白,采用玻璃珠法破菌,提取蛋白質(zhì)后通過SDS-PAGE凝膠電泳鑒定表達產(chǎn)物。經(jīng)響應(yīng)面優(yōu)化后,在37℃培養(yǎng)工程菌至菌液OD600=0.74后,加入異丙基-β-D-硫代半乳糖苷(IPTG)至終濃度為0.40 mmol/L,在25.51℃條件下誘導(dǎo)培養(yǎng)8 h,可獲得最高產(chǎn)量的可溶性目的蛋白,為128μg/m L。最優(yōu)表達條件的獲得,為后期純化該可溶性蛋白和鑒定Fa MYB5蛋白與順勢作用元件的作用情況提供了一定的理論依據(jù)。
[Abstract]:The optimal conditions for prokaryotic expression of pET-32a (-Fa MYB5) fusion protein were investigated by central combination design. Four factors were investigated under different induction temperature, induction time, initial concentration of bacterial solution and final concentration of isopropyl- 尾 -Dthiogalactoside (IPTG). The fusion protein of pET-32a (-Fa) MYB5 was expressed in Escherichia coli with soluble protein content as the index. The protein was extracted by glass bead method, and the expressed product was identified by SDS-PAGE gel electrophoresis. The expression product was optimized by response surface optimization. After the engineering bacteria were cultured to OD600=0.74 solution at 37 鈩，

本文編號：1682241