MIDSCI™ Cryo Label Sheets

ig BL21 (DE3)
Intact Genomics chemically competent BL21(DE3) E. coli cells are suitable for transformation and routine protein expression.

Specifications
Competent cell type:                Chemically competent
Derivative of:                            BL21(DE3)
Species:                                   E. coli
Format:                                    Tubes
Transformation efficiency:         1.0 x 109 cfu/µg pUC19 DNA
Blue/white screening:               Yes
Shipping condition:                   Dry ice

Quality Control
Quality control is performed following the production of each new lot of product to ensure that it meets the quality standards and specifications designated for the product. Each lot is repeatedly compared side-by-side with leading competitors to ensure our products outperform the competitor before product launching. 

Reagents Needed for One Reaction
BL21(DE3) Chemically Competent Cells:             50 µl
DNA (or pUC19 Control, 10 pg/µl):                         1 µl
Recovery Medium:                                                 1 ml

Contents & Storage
BL21(DE3) Competent Cells:                -80 °C
pUC19 Control DNA:                            -20 °C
Recovery Medium:                                  4 °C

Genomic Features
BL21(DE3) chemically competent cells have the following features:

• Widely used host background
• T7 Expression Strain
• Deficient in both lon (1) and ompT proteases
• Resistant to phage T1 (fhuA2)
• B Strain

Genotype
F-ompT hsdS(rB- mB-) gal dcm (DE3)

General Guidelines
Follow these guidelines when using BL21(DE3) chemically competent E. coli.

• Handle competent cells gently as they are highly sensitive to changes in temperature or mechanical lysis caused by pipetting.
• Thaw competent cells on ice, and transform cells immediately following thawing. After adding DNA, mix by tapping the tube gently. Do not mix cells by pipetting or vortexing.

Transformation Protocol

• Remove competent cells from the -80 °C freezer and thaw completely on wet ice (10-15 minutes).
• Aliquot 1-5 µl (1 pg-100 ng) of DNA to the chilled microcentrifuge tubes on ice.
• When the cells are thawed, add 50 μl of cells to each DNA tube on ice and mix gently by tapping 4-5 times. For the pUC19 control, add 1 µl of (10 pg/µl)
• DNA to a chilled microcentrifuge tube, prior to adding 50 µl of cells. Mix well by tapping.
• Incubate the cells with DNA on ice for 30 minutes.
• After 30 minute ice incubation, heat shock the cells at 42 °C for 45 seconds.
• Transfer the tubes to ice for 2 minutes.
• Add 950 µl of Recovery Medium or any other medium of choice to each tube.
• Incubate tubes at 37 °C for 1 hour at 210 rpm.
• Spread 50 μl to 200 μl from each transformation on pre-warmed selection plates.
• Incubate the plates overnight at 37 °C.