ig 10B Chemically Competent Cells

Description

Intact Genomics ig™ 10B chemically competent cells (E. coli) are suitable for high efficiency transformation in a wide variety of applications such as cloning and sub-cloning.  This derivative of DH10B provides the highest efficiency in the industry.

Specifications

Competent cell type:                Chemically competent
Derivative of:                            DH10B™
Species:                                   E. coli
Format:                                    Tubes
Transformation efficiency:         ≥1.0 x 10¹⁰ cfu/µg pUC19 DNA
Blue/white screening:               Yes
Shipping condition:                   Dry ice

Reagents Needed for One Reaction

ig™ 10B chemically competent cells:     50 µl
DNA (or pUC19 Control, 10 pg/µl):          1 µl
Recovery medium:                                  1 ml

Product Includes & Storage

1. ig™  10B competent cells:    -80 ºC
2. pUC19 control DNA:             -20 ºC
3. Recovery medium:                  4 ºC

Genomic Features

ig™ 10B (DH10B derivative) chemically competent cells have the following features:

• Φ80lacZΔM15 marker provides α-complementation of the β-galactosidase gene with blue/white screening
• mcrA genotypic marker and the mcrBC, mrr deletion allows for cloning DNA that contains methylcytosine and methyladenine 

Genotype

F – mcrA ∆(mrr-hsdRMS-mcrBC) endA1 recA1 φ80dlacZ∆M15 ∆lacX74 araD139 ∆(ara, leu)7697 galU galK rpsL (StrR) nupG λ-

Quality Control

Transformation efficiency is tested by using the pUC19 control DNA supplied with the kit and the high efficiency transformation protocol listed below. Transformation efficiency should be ≥1 x 10¹⁰ CFU/µg pUC19 DNA.

Untransformed cells are tested for appropriate antibiotic sensitivity.

General Guidelines

Follow these guidelines when using ig™ 10B 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.

Calculation of Transformation Efficiency
Transformation Efficiency (TE) is defined as the number of colony forming units (cfu) produced by transforming 1µg of plasmid into a given volume of competent cells.

TE = Colonies/µg/Dilution

Transform 1 µl of (10 pg/µl) pUC19 control plasmid into 50 µl of cells, add 950 µl of Recovery Medium. Dilute 10 µl of this in 990 µl of Recovery Medium and plate 50 µl. Count the colonies on the plate the next day. If you count 100 colonies, the TE is calculated as follows:

Colonies = 100
µg of DNA = 0.00001
Dilution = 50/1000 x 10/1000 = 0.0005
TE = 100/.00001/.0005 = 2.0×10¹⁰1011-06 1011-12 1011-24 1012-12 1012-24 1012-48 1014-24 1014-48 1018-96