大肠杆菌表达载体：pTWIN1 百欧博伟生物
北京百欧博伟生物技术有限公司（biobw）是北京的一家专业于生物技术的研究与广泛运用及推广的高科技公司，位于北京企业林立的丰台区造甲街，生物技术推广及运用早于2011年开始，成立公司于2013年一月。本公司主要提供色谱耗材、色谱仪器、固相萃取装置、化学试剂、样品瓶、氘灯、标准品、微生物技术、菌种保藏服务以及ATCC产品代理等产品及服务。
基本信息
平台编号：bio-107412
质粒类型：大肠杆菌表达载体
克隆方法：多克隆位点，限制性内切酶
载体大小：7375bp
载体抗性：Ampicillin (氨苄青霉素)
载体描述
pTWIN1 is an E. coli plasmid cloning vector designed for recombinant protein expression, labeling, and cyclization using the IMPACT-TWIN Kit (NEB #E6901) (1). It contains the pMB1 origin of replication from pBR322 and is maintained at a similar copy number to pBR322; in addition, pTWIN1 also contains an M13 origin of replication.
The multiple cloning site (MCS) is positioned to allow translational fusion of an intein tag to the N-terminus, C-terminus, or both, of the cloned target protein. The mini-inteins encoded by pTWIN1, the Ssp DnaB intein and the Mxe GyrA intein, cleave the peptide bond at their C- and N-termini, respectively (1-4). The chitin binding domain (CBD) from B. circulans, fused to each intein, facilitates purification of the intein-target protein precursor.
Transcription of the intein tags is controlled by the inducible T7 promoter, requiring E. coli strains containing integrated copies of the T7 RNA polymerase gene [e.g., NEB #C2566, #C2833 or BL21(DE3)] for expression. Basal expression from the T7 promoter is minimized by the binding of the Lac repressor, encoded by the lacI gene, to the lac operator immediately downstream of the T7 promoter (5). Translation of the intein tags utilizes the translation initiation signal (Shine Dalgarno sequence) from the strongly expressed T7 gene 10 protein (φ10).
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