生物技术通讯
生物技術通訊
생물기술통신
LETTERS IN BIOTECHNOLOGY
2014年
1期
45-48
,共4页
李强%刘先凯%冯尔玲%朱力%王沛%王红%杨新华%赵美玲%姜永强%王恒樑
李彊%劉先凱%馮爾玲%硃力%王沛%王紅%楊新華%趙美玲%薑永彊%王恆樑
리강%류선개%풍이령%주력%왕패%왕홍%양신화%조미령%강영강%왕항량
炭疽芽胞杆菌%FtsE蛋白%融合表达
炭疽芽胞桿菌%FtsE蛋白%融閤錶達
탄저아포간균%FtsE단백%융합표체
Bacillus anthracis%FtsE protein%fusion expression
目的：构建炭疽芽胞杆菌FtsE蛋白的原核表达载体，实现其在原核表达系统中的可溶性表达，并纯化融合蛋白。方法：用PCR方法从炭疽芽胞杆菌A16R株扩增得到ftsE基因片段，酶切后连接到pET28a原核表达载体，构建重组表达质粒pET28a-ftsE，转化大肠杆菌BL21（DE3）菌株，筛选可溶性诱导表达与纯化融合蛋白的条件，以获得高纯度融合蛋白。结果：构建了FtsE蛋白的融合表达载体，并在大肠杆菌中获得高效表达；在20℃下，经0.1 mmol/L IPTG诱导3 h表达的产物主要是可溶性蛋白，经Ni-NTA亲和层析纯化获得了高纯度的FtsE融合蛋白，经Western印迹检测，目的蛋白表达正确。结论：实现了炭疽芽胞杆菌FtsE蛋白原核表达系统的可溶性表达并获得了高纯度融合蛋白，为后续研究奠定了基础。
目的：構建炭疽芽胞桿菌FtsE蛋白的原覈錶達載體，實現其在原覈錶達繫統中的可溶性錶達，併純化融閤蛋白。方法：用PCR方法從炭疽芽胞桿菌A16R株擴增得到ftsE基因片段，酶切後連接到pET28a原覈錶達載體，構建重組錶達質粒pET28a-ftsE，轉化大腸桿菌BL21（DE3）菌株，篩選可溶性誘導錶達與純化融閤蛋白的條件，以穫得高純度融閤蛋白。結果：構建瞭FtsE蛋白的融閤錶達載體，併在大腸桿菌中穫得高效錶達；在20℃下，經0.1 mmol/L IPTG誘導3 h錶達的產物主要是可溶性蛋白，經Ni-NTA親和層析純化穫得瞭高純度的FtsE融閤蛋白，經Western印跡檢測，目的蛋白錶達正確。結論：實現瞭炭疽芽胞桿菌FtsE蛋白原覈錶達繫統的可溶性錶達併穫得瞭高純度融閤蛋白，為後續研究奠定瞭基礎。
목적：구건탄저아포간균FtsE단백적원핵표체재체，실현기재원핵표체계통중적가용성표체，병순화융합단백。방법：용PCR방법종탄저아포간균A16R주확증득도ftsE기인편단，매절후련접도pET28a원핵표체재체，구건중조표체질립pET28a-ftsE，전화대장간균BL21（DE3）균주，사선가용성유도표체여순화융합단백적조건，이획득고순도융합단백。결과：구건료FtsE단백적융합표체재체，병재대장간균중획득고효표체；재20℃하，경0.1 mmol/L IPTG유도3 h표체적산물주요시가용성단백，경Ni-NTA친화층석순화획득료고순도적FtsE융합단백，경Western인적검측，목적단백표체정학。결론：실현료탄저아포간균FtsE단백원핵표체계통적가용성표체병획득료고순도융합단백，위후속연구전정료기출。
Objective: To construct ftsE recombinant expression plasmid and optimize the expression conditions, and to purify the fusion protein. Methods: The amplified ftsE gene fragment by PCR was inserted into pET28a to construct fusion protein expression plasmid. Then the recombinant plasmid was transformed into E.coli BL21(DE3), and the conditions of induction and purification of FtsE fusion protein were optimized. Results: The pET28a-ftsE recombinant plasmid was constructed successfully. Soluble fusion protein was obtained by induction of 0.1 mmol/L IPTG at 20℃ lasted 3 hours. And the FtsE fusion protein was purified by Ni-NTA affinity chromatography and verified by Western blot. Conclusion: Soluble FtsE fusion protein was achieved, and high purity of the FtsE fu-sion protein was obtained. This work may lay a foundation for further study of the functions of FtsE protein.
