Product Class: Other

T7 Express lysY Competent E. coli (High Efficiency)

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Product Introduction

High efficiency chemically competent E. coli cells suitable for T7 expression with enhanced reduction of basal expression.

  • T7 RNA Polymerase in the lac operon - no λ prophage
  • Control of T7 RNA Polymerase by lysozyme allows potentially toxic genes to be expressed
  • Deficient in proteases Lon and OmpT
  • Resistant to phage T1 (fhuA2)
  • Free of animal products
  • No dry ice surcharge on NEB competent cell shipments
Catalog # Size Concentration
C3010I 6.0 x 0.2 ml

Product Information

Description

Highlights

  • Free of animal products
  • Control of T7 RNA Polymerase by lysozyme allows potentially toxic genes to be expressed
  • LysY is a variant of T7 lysozyme lacking amidase activity, thus cells are less susceptible to lysis during induction
  • Maintenance of lysozyme plasmid does not require antibiotic selection
  • Enhanced BL21 derivative
  • Does not restrict methylated DNA (McrA-, McrBC-, EcoBr-m-, Mrr-)
  • Deficient in proteases Lon and OmpT
  • Lysozyme for expression control
  • T7 RNA Polymerase in the lac operon - no λ prophage
  • No Cam requirement
  • Fast growth from colonies
  • Lysozyme on miniF enhances stability
  • Resistant to phage T1 (fhuA2) Cam, Nit
  • B Strain

Transformation Efficiency

0.6-1 x 109 cfu/μg pUC19 DNA Enhanced BL21 derivative, chemically competent E. coli cells suitable for high efficiency transformation and protein expression.

Genotype

MiniF lysY (CamR) / fhuA2 lacZ::T7 gene1 [lon] ompT gal sulA11 R(mcr-73::miniTn10--TetS)2 [dcm] R(zgb-210::Tn10--TetS) endA1 ∆(mcrC-mrr)114::IS10
This product is related to the following categories:
T7 Expression,
E. coli Protein Expression Strains Products,
Protein Expression Products,
E. coli Expression Strains Products,
This product can be used in the following applications:
T7 Expression,
Toxic Protein Expression,
Peptide Ligation,
Expression of Difficult Proteins,
Protein Expression in E. Coli,
Protein Expression

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • C3010I     -80    
      T7 Express lysY Competent E. coli (High Efficiency) C3010IVIAL -80 6 x 0.2 ml Not Applicable

Properties & Usage

Antibiotic for Plasmid Selection

Antibiotics for Plasmid Selection Working Concentration
Ampicillin 100 µg/ml
Carbenicillin 100 µg/ml
Kanamycin 30 µg/ml
Streptomycin 25 µg/ml
Tetracycline 15 µg/ml

Shipping Notes

  • Ships on dry ice

Antibiotic Resistance

  • nit
  • cam

Features

  • T7 expression
  • Protease deficient
  • Better reduction of basal expression

Application Features


Transformation of a toxic mammalian clone into E. coli hostsTransformation of a toxic mammalian clone into E. coli hosts
 A T7 expression plasmid and the same plasmid containing a gene encoding a toxic mammalian protein were transformed into each host. Comparison of the relative transformation efficiencies demonstrates that the T7 Express hosts provide the levels of control necessary for transformation of potentially toxic clones. BL21(DE3) could not be transformed with the toxic clone.
T7-controlled expression of a non-toxic protein in E. coli hostsT7-controlled expression of a non-toxic protein in E. coli hosts
A T7 expression plasmid containing a gene encoding an E. coli protein was transformed into each host, grown to 0.6 OD and induced for 3 hours. Comparison of soluble extracts from uninduced (-) and induced (+) cells shows superior control of basal expression in the T7 Express hosts while maintaining high levels of induced expression.
Effect of heat shock time on T7 Express lysY competent E.coli transformation efficiencyEffect of heat shock time on T7 Express lysY competent E.coli transformation efficiency
50 μl of competent cells were transformed with 100 pg of pUC19 control DNA following the provided High Efficiency Transformation Protocol except heat shock time varied from 0 to 80 seconds.
Effect of DNA incubation time on T7 Express lysY competent E.coli  transformation efficiencyEffect of DNA incubation time on T7 Express lysY competent E.coli transformation efficiency
50 μl of competent cells were transformed with 100 pg of pUC19 control DNA following the provided High Efficiency Transformation Protocol except DNA incubation time varied from 0 to 40 minutes.

Product Notes

  1. Maintenance of the miniF plasmid does not require antibiotic selection. If chloramphenicol is added, use 10 µg/ml final concentration.
  2. CAUTION: This product contains DMSO, a hazardous material. Review the MSDS before handling.
  3. STORAGE AND HANDLING: Competent cells should be stored at -80°C. Storage at -20°C will result in a significant decrease in transformation efficiency. Cells lose efficiency whenever they are warmed above -80°C, even if they do not thaw.

Protocols, Manuals & Usage

Protocols

  1. High Efficiency Transformation Protocol (C3010)
  2. 5 Minute Transformation Protocol (C3010)
  3. Protocol for Expression Using T7 Express lysY (C3010)

Usage & Guidelines

Application Notes

Tools & Resources

Selection Charts

FAQs & Troubleshooting

FAQs

  1. Why are there no colonies or no growth in liquid culture (C3010)?
  2. Why is there no protein visible on gel or no activity (C3010)?
  3. Why is induced protein insoluble (C3010)?
  4. What are the solutions/recipes (C3010)?
  5. What are the strain properties (C3010)?
  6. Can I store competent cells at -20°C instead of -80°C?
  7. What is the difference between NEB #C3010H and NEB #C3010I?
  8. What is the optimal heat shock time for this strain (NEB #C3010H and NEB #C3010I)?
  9. How long should I incubate cells on ice after DNA has been added (NEB #C3010H and NEB #C3010I)?
  10. Is T7 Express lysY (NEB #C3010H and NEB #C3010I) compatible with auto-induction procedures?
  11. Which kind of transformation tubes should be used?
  12. What volume of DNA can be added into competent cells?
  13. What s the shelf life for this strain (NEB #C3010H and NEB #C3010I)?
  14. Are NEB's competent cells compatible with the “Mix & Go" protocol?
  15. Is antibiotic selection necessary to maintain the MiniF-lysY plasmid?
  16. Is T7 expression subject to catabolite repression in NEB T7 Express or SHuffle strains?

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email [email protected].

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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