Express® E. coli Lysis Reagent  |NEB酶试剂 New England Biolabs

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产品信息

NEBExpress® E. coli Lysis Reagent is a chemical lysis solution composed of a proprietary mix of non-ionic and zwitterionic detergents and Tris-based buffer. It allows disruption of E. coli cells without denaturing soluble proteins.

Figure 1: Optimal protein extraction with NEBExpress T4 Lysozyme in the presence of NEBExpress E. coli Lysis Reagent.

Express® E. coli Lysis Reagent  |

T7 Express E. coli  expressing vGFP was lysed with NEBExpress T4 Lysozyme (T4L) in 50 mM Tris-HCl pH 7.5 or in NEBExpress E. coli Lysis Reagent (NEB #P8116S): 4 UOD600 of cell pellet were resuspended in 200 µL of Tris buffer or NEBExpress E. coli Lysis Reagent and lysed for 5 minutes at room temperature in the presence of NEBExpress T4 Lysozyme at 0, 0.1 or 1 ug per 1 mL of cell suspension. The soluble proteins were harvested by centrifugation and analyzed on SDS-PAGE.

产品类别:
Competent Cells Products

应用:
High-throughput cloning and automation solutions,
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
    • P8116S     25    
        NEBExpress® E. coli Lysis Reagent P8116SVIAL 25 1 x 100 ml 1 X
    • P8116L     25    
        NEBExpress® E. coli Lysis Reagent P8116SVIAL 25 5 x 100 ml 1 X

  • 优势和特性

    Features

    • Scalable lysis reactions from small to large bacterial cell pellets and compatible with high throughput workflows
    • Suitable for extracting proteins, especially thermosensitive proteins vulnerable to mechanical lysis procedures
    • Fast and simple protocol: harvested cells are resuspended and briefly incubated in the Lysis Reagent to extract soluble proteins. Insoluble cell debris is removed by centrifugation, and the clarified extract is ready to use and compatible with affinity resins.
    • Compatible with analyses such as SDS-PAGE, Western blots, activity assays, immunoprecipitation, and downstream purification 
    • Suitable for disrupting most Gram-negative bacterial cells
    • Small (100 ml) and large kits (5 x 100 ml) are suitable for lysing 20 to 100 grams of cells, respectively.
     

  • 相关产品

    相关产品

    • m0303-dnase-i-rnase-free
    • T7 表达 lysY E. coli 感受态细胞（高效级）
    • BL21(DE3) E. coli 感受态细胞
    • T7 表达 E. coli 感受态细胞（高效级）
    • Amylose Resin
    • Amylose Resin High Flow
    • 几丁质树脂
    • NEBExpress® Ni-NTA 磁珠
    • NEBExpress® Ni Spin Columns
    • NEBExpress® Ni Resin
    • E8201 NEBExpress MBP 融合表达及纯化系统
    • IMPACT™ Kit
    • 亲水性链霉亲和素磁珠
    • SNAP-捕获磁珠
    • Color Prestained Protein Standard, Broad Range (10-250 kDa)
    • 蓝色蛋白上样染料
    • NEBExpress® T4 Lysozyme
    • 微球菌核酸酶

  • 注意事项

    1. NEBExpress E. coli Lysis Reagent is compatible with: 

      • Tris, phosphate, and PIPPS-based buffers ranging from pH 5.5-10.
      • EDTA. To prevent interference with metal affinity resins use < 0.1-0.5 mM EDTA. If using NEBExpress Ni Resin (NEB #S1428S) or Ni Spin Columns (NEB #S1427S/L), there is no limitation on EDTA use, as the resin is chemically resistant to EDTA chelation.
      • < 1 M NaCl (precipitation will occur at or above 1 M NaCl).
      • Protease inhibitors 
      • Reducing agent such as DTT, β-mercaptoethanol, TCEP, THP. To prevent interference with metal affinity resins, use 1-2 mM β-mercaptoethanol or 1-5 mM TCEP. NEBExpress Ni Resin (NEB #S1428) and Ni Spin Columns (NEB #S1427S/L) are both chemically resistant to these reducing agents. 
      • BCA Protein assay
      • SDS-PAGE and Western blots
      • 1 µL NEBExpress® T4 Lysozyme (NEB# P8115) per 1 mL of lysate for improved protein extraction efficiency. The lysate may become viscous due to the release of DNA. For optimal lysis yield, add 1mM CaCl2 and 1 µL of Micrococcal Nuclease (NEB #M0247) per 1 mL of lysate, mix and incubate for 5 minutes at room temperature.
       
    2. NEBExpress E. coli Lysis Reagent can be added directly to frozen cell pellets; however, allow this mixture to reach room temperature as the lysis will be more efficient.
    3. Use 0.025 – 0.075 mL NEBExpress E. coli Lysis Reagent per unit OD600 (UOD600) cells harvested. For example, for a 5 mL culture harvested at OD600 of 1, 5 UOD600 of cells are obtained (5 mL x 1 OD600 = 5 UOD600), thus requiring 0.125 to 0.375 mL of Lysis Reagent. 
    4. If weighing the cell pellet, use 5 mL NEBExpress E. coli Lysis Reagent per 1 gram of cell paste. Note that cells resuspended in Lysis Reagent at higher cell densities will not lyse as efficiently.
    5. Extraction efficiency is strain dependent. Efficiency is highest with BL21 strains, its derivatives with DE3 or lysY, E. coli K-12 strains, and various other Gram-negative bacteria (e.g., Salmonella, Vibrio, Pseudomonas).
    6. Storing NEBExpress E. coli Lysis Reagent at 4°C or below may cause precipitation of the detergents. However, the lysis efficiency of the reagent can be fully restored by incubation at room temperature with gentle mixing to dissolve any precipitate.

操作说明、说明书 & 用法

  • 操作说明

    1. Protocol for cell lysis using NEBExpress E. coli Lysis Reagent (NEB #P8116)

FAQs & 问题解决指南

  • FAQs

    1. How much NEBExpress® E. coli Lysis Reagent should I use?
    2. Why do I see precipitation in the NEBExpress® E. coli Lysis Reagent?
    3. Can I use NEBExpress® E. coli Lysis Reagent that has been frozen?
    4. Should I chill the NEBExpress® E. coli Lysis Reagent prior to use?
    5. Should I freeze the cell pellets prior to lysing with NEBExpress® E. coli Lysis Reagent?
    6. What buffer is present in the NEBExpress® E. coli Lysis Reagent?
    7. Can I supplement the NEBExpress® E. coli Lysis Reagent with NaCl?
    8. Can I change the pH of NEBExpress® E. coli Lysis Reagent?
    9. Is the NEBExpress® E. coli Lysis Reagent compatible with EDTA?
    10. Can I add a protease inhibitor to the NEBExpress® E. coli Lysis Reagent reaction?
    11. Do I need to add lysozyme to aid in lysis?
    12. My lysate is viscous; how can I resolve this?
    13. Can I use a reducing agent with NEBExpress® E. coli Lysis Reagent?
    14. Does NEBExpress® E. coli Lysis Reagent lyse gram-positive bacterial cells or yeast cells?
    15. Does NEBExpress® E. coli Lysis Reagent lyse gram-negative bacterial cells?
    16. My protein forms inclusion bodies. Can I still use this reagent to lyse my cells?
    17. Can I add NEBExpress® E. coli Lysis Reagent directly to my cell culture to lyse the cells?

  • 问题解决指南

    • Troubleshooting Guide for NEBExpress E. coli Lysis Reagent (NEB # P8116)