pMAL-c6T Vector |NEB酶试剂 New England Biolabs

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产品信息

The pMAL-c6T Vector provides a method for producing a protein expressed from a cloned gene or open reading frame. The cloned gene is inserted downstream from the malE gene of E. coli, which encodes maltose-binding protein (MBP), resulting in the expression of an MBP fusion protein in the cytoplasm (1,2). The MBP has been engineered for tighter binding to amylose resin. The method uses the strong “tac” promoter and the malE translation initiation signals to give high-level expression of the cloned sequences (3,4), and a one-step purification of the fusion protein using MBP’s affinity for maltose (5). The vector expresses an N-terminal hexahistidine tagged malE gene (lacking its secretory signal sequence) followed by a multiple cloning site containing a TEV protease recognition sequence and stop codons in all three frames. This allows MBP to be cleaved from the protein of interest by TEV Protease after purification. The vector also carries the lacIq gene, which encodes for the Lac repressor. This keeps expression from Ptac low in the absence of IPTG induction. 

Source: NEB-10 beta competent E. coli (pMAL-c6T)

Figure 1: pMAL-c6T Vector

pMAL-c6T Vector |

The pMAL- c6T Vector has an exact deletion of the malE signal sequence. Arrows indicate the direction of transcription. Unique restriction sites in the multiple cloning site (MCS) are indicated.

产品来源

NEB-10 beta competent E. coli (pMAL-c6T)

产品类别:
NEBExpress MBP Fusion and Purification System,
Bacterial E. coli Protein Expression Products,
DNA Plasmids & Substrates Products,

Protein Expression Products

应用:
Target Protein Insolubility ,
Non-T7 Expression,
Expression of Difficult Proteins,

Protein Expression in E. Coli,

Protein Expression

  • 产品组分信息

    本产品提供以下试剂或组分：

    NEB # 名称 组分货号 储存温度 数量 浓度
    • N0378S     -20    
        pMAL-c6T Vector N0378SVIAL -20 1 x 10 µg 200 µg/ml

  • 特性和用法

    亲和标签

    Maltose-Binding Protein (MBP)

    序列文件

    Fasta GenBank

  • 优势和特性

    Features

    • The recommended vector for use with the NEBExpress® MBP Fusion and Purification System (NEB #E8201)
    • Fusion to MBP significantly enhances proper folding and solubility of target proteins
    • Vector includes His-tag and TEV Protease recognition site
    • Two-step purification: amylose resin elution followed by TEV Protease cleavage and Ni resin isolation results in a highly pure, tag-free target protein

  • 相关产品

    相关产品

    • E8201 NEBExpress MBP 融合表达及纯化系统
    • Amylose Resin
    • TEV Protease
    • Anti-MBP Monoclonal Antibody
    • NEB® 10-beta E. coli 感受态细胞（高效级）
    • c2523-neb-express-competent-e-coli-high-efficiency

  • 注意事项

    1. A detailed product manual for the NEBExpress MBP Fusion and Purification System with protocols can be found here.
    2. NEB 10-beta Competent E. coli (High Efficiency) (NEB #C3019) is recommended for propagation and subcloning. NEB Express Competent E. coli (High Efficiency) (NEB #C2523) is recommended for expression using this vector.
    3. This vector conveys ampicillin resistance for propagation in E. coli.

  • 参考文献

    1. Guan, C. et al (1987). Gene. 67, 21–30.
    2. Maina, C.V. et al (1988). Gene. 74, 365–373.
    3. Amann, E. et al (1985). Gene. 40, 183–190..
    4. Duplay, P. et al (1984). J. Biol. Chem. 259, 10606–10613.
    5. Kellerman, O.K. et al (1982). Methods in Enzymology. 90, 459–463.

操作说明、说明书 & 用法

  • 操作说明

    1. NEBExpress MBP Fusion and Purification System Quick Start Protocol (NEB #E8201)
    2. Cloning a PCR Fragment Into a pMAL Expression Vector (E8201)
    3. NEBuilder Assembly of a PCR Fragment
    4. Transformation Protocol (NEB #E8201)
    5. Small Scale Affinity Chromatography (NEB #E8201)
    6. Large Scale Affinity Chromatography (NEB #E8201)
    7. Cleavage of the Fusion Protein
    8. Separating the Protein of Interest from MBP after TEV Protease Cleavage (NEB #E8201)

  • 说明书

    产品说明书包含产品使用的详细信息、产品配方和质控分析。

    • manualE8201

工具 & 资源

  • 选择指南

    • Protein Expression and Purification Selection Chart

  • Web 工具

    • DNA Sequences and Maps Tool

FAQs & 问题解决指南

  • FAQs

    1. What strain(s) do you recommend as hosts for the pMAL vectors?
    2. What primers should I use to sequence the ends of my insert after I clone it into a pMAL vector?
    3. What is the minimum size of a fragment that can be cloned into pMAL and expressed fused to MBP? Can short peptide sequences (~ 10 amino acids) be added onto MBP?
    4. What is the DNA sequence of this product?
    5. What are the differences between the generations of pMAL vectors (e.g. pMAL-c2 vs. pMAL-c5 vs. pMAL-c6 or pMAL-p2 vs. pMAL-p5)?