Zr viral dna kit™

Premade Z-CompetentTM E. coli Cells

Highlights
• Feature fast transformation kinetics: No heat shock, no lengthy incubations, no outgrowth • High transformation efficiencies: 108-109 transformants/µg plasmid DNA. • Simple: Mix DNA with cells for a few seconds and plate. Mix & Go! Contents
Product Contents and Description ……….…….1 Protocol………………………………………….2, 3 Appendix.………………………………………….3 Ordering Information.…………………………….4 List of Related Products.………………….……5 Toll Free: 1-888-882-9682 • Fax: 1-714-288-9643 • Web: www.zymoresearch.com • E-mail: [email protected] Product Contents
be dissatisfied with this product please call 1-888- Z-Competent™ E. coli Cells
Instruction Manual
Note – Chemically Z-Competent™ cells are stable for 6 months at < -70oC. Reagents are routinely tested on a lot-to-lot
basis to ensure they provide maximal performance and reliability. Product Description

Z-Competent™ E. coli are premade chemically competent cells used for simple
and highly efficient DNA transformation. Z-Competent™ E. coli cells are made chemically competent by a unique method using ZymoBroth™ that completely
eliminates the need for heat shock and related procedures. For transformation, DNA can be added directly to Z-Competent™ cells and the mixture spread directly to a culture plate - Mix & Go! Transformation efficiencies typically range from 108–109 transformants/µg of pUC19 DNA (see figures below), which make the cells optimal for cloning, sub-cloning, library construction, etc. Premade Z-Competent™ cells are supplied as a pack of 10 convenient 100 µl/tube aliquots or in a 96-well format (12x8- Z-Competent™ E. coli cells prepared with ZymoBroth™ display fast transformation kinetics and high
transformation efficiencies. Figures above show the transformation kinetics for Z-Competent™ JM109 and BL21
strains of E. coli generated using ZymoBroth™ and SOB growth media. Plasmid DNA (pUC19) was used for transformation and the data are the averages of three individual experiments. Note -TM Trademarks of Zymo Research Corporation. This product is for research use only and should only be used by trained professionals. Wear protective gloves and eye protection. Fol ow the safety guidelines and rules enacted by your research institution or facility. Toll Free: 1-888-882-9682 • Fax: 1-714-288-9643 • Web: www.zymoresearch.com • E-mail: [email protected] Protocol

For Technical Assistance,
Pre-warm culture plates to 37oC before starting. please contact Zymo
Research’s Technical
Department at 1-888-882-
Standard Transformation Procedure
9682 or E-mail to [email protected]. Single Tube Aliquots
1. Add 1-5 µl plasmid DNA to a tube of thawed Z-Competent™ cells on ice, mix gently for a few seconds (try to keep the added volume of DNA less than 5% of 2. Incubate on ice for 2-5 minutes (maximum 60 minutes). 3. Spread 50-100 µl onto a pre-warmed culture plate. Incubate the plate at the appropriate temperature (e.g., 37°C) for the colonies to grow. 96-Well Format (8-Well Strips)
Thaw tube strips of frozen Z-Competent™ cells on ice (or 0-2°C). 1. Add 1-3 µl DNA to each tube of thawed Z-Competent™ cells and mix gently by 2. Incubate on ice (or 0-2°C) for 2-5 minutes (maximum 60 minutes). 3. Spread 25-50 µl of the mixtures onto pre-warmed culture plates. 4. Incubate the plate at 37°C or the appropriate temperature for the colonies to
Note: The procedures above are for plasmids containing Ampicillin resistant markers. If Kanamycin, Tetracycline,
Chloramphenicol, Erythromycin or any non-lactamase selection markers are used, an outgrowth step is required
prior to plating. (see Notes Section 4 on page 3 regarding Outgrowth)
Important! Since chemically competent cells are extremely sensitive to changes in temperature,
transformation should be performed immediately after thawing. The cells should avoid being
exposed to room temperature for more than a few seconds. To mix cells after DNA addition,
gently tap the tube with your fingers and then shake the tube downwards in a single motion from
the elbow to collect the mixture at the bottom of the tube. Avoid repeated pipetting. Immediately
transfer the transformation mixture(s) to ice or an ice bath (0oC).
Fast Transformation of Z-Competent™ Cells* - Mix & Go!
1. Add 1-5 µl plasmid DNA to a tube of thawed Z-Competent™ cells on ice, mix gently for a few seconds (try to keep the added volume of DNA less than 5% of the total). 2. Spread 50-100 µl of the mixture onto a pre-warmed (37°C) culture plate containing Ampicillin. Incubate the plate at the appropriate temperature (e.g., 37°C) for the colonies to grow. *For Ampicillin selection only. For selection with other antibiotics, see Notes Section 4 on page 3. Toll Free: 1-888-882-9682 • Fax: 1-714-288-9643 • Web: www.zymoresearch.com • E-mail: [email protected] Notes for High Efficiency Transformation
1. E. coli Strains
Different E. coli strains vary in their ability to be transformed with DNA. Strains like JM109, C600, TG1, and DH5a* typically yield the highest transformation efficiencies. 2. Incubation Time
The “Mix & Go!” procedure (page 2) will work for most transformations using Ampicillin selection and not requiring outgrowth (see Section 4 below). The highest transformation efficiencies can be obtained by incubating Z-Competent™ cells with DNA on ice for 2-5 minutes (60 minutes maximum) prior to plating. 3. Prewarming Culture Plates
Chilled plates will decrease Z-Competent™ cell transformation efficiency. It is recommended that culture plates be pre-warmed to >20°C (preferably 37°C) prior to 4. Addition of SOC Medium to Transformation Mixtures (Outgrowth)
When selecting with Kanamycin, Tetracycline, etc., an outgrowth performed in SOC medium is required for efficient transformation. In most cases, this step can be omitted when selecting with Ampicillin. After the transformation mixture has incubated on ice for 5-10 min, add 4 volumes of SOC (400 µl of SOC to 100 µl of transformation mixture) and incubate for 1 hour at 37 300 rpm. Afterwards, spread the mixture directly onto pre-warmed culture plates. Reducing agents [e.g., DTT (Dithiothreitol) and 2-ME (β-mercaptoethanol)] are not Appendix
SOB Recipe: (1 Liter)
Adjust pH to 6.0-7.0 with NaOH. Autoclave at 10 psi for 15-20 minutes. SOC Recipe: (100 ml)
Add 1 ml of a 2 M filter-sterilized glucose solution or 2 ml of 20% (w/v) glucose solution to 100 LB Agar (1 Liter)
Adjust the pH to 7.0 with 5 N NaOH. Autoclave at 15 psi for 15-20 minutes. Toll Free: 1-888-882-9682 • Fax: 1-714-288-9643 • Web: www.zymoresearch.com • E-mail: [email protected] Genotypes

JM109
F`traD36 lacIq Δ(lacZ)M15 pro A+ B+ / e14- (McrA- ) Δ(lac-proAB)thi gyrA96 (NaIr )endA1 hsdR17(r -
References:
1.) Sheridan, P. et al. Phylogenetic
Comments: Partly restriction-deficient; good strain for cloning repetitive DNA(recA- ). Suppresses many amber mutations
Analysis of Anaerobic
when glutamine is available but not the S Psychrophilic Enrichment Cultures
100 or S7 mutation of λ, e.g., λgt11. Can be used for M13 cloning/sequencing and Obtained from a Greenland Glacier
Ice Core, Appl. Envir. Microbiol., Apr
2.) Yokobayashi, Y. et al. From the
k ,mk ) gyrA96 relA1 thi mcrA Δ(lac-proAB) ΔaraB:: ΔR, cat F’[traD36 proAB+ lacIq lacZ Cover: Directed evolution of a
genetic circuit, PNAS, Dec 2002; 99:
16587 – 16591.
Comments: Includes chromosomally encoded bacteriophage lambda R gene. Partly restriction-deficient; good strain for
cloning repetitive DNA(recA- ). Suppresses many amber mutations when glutamine is acceptable but not the S100 or S7 3.) Trent, J. et al. A Ubiquitously
mutation of λ, e.g., λgt11. Can be used for M13 cloning/sequencing and blue/white screening. Expressed Human Hexacoordinate
Hemoglobin, J. Biol. Chem., May
XJa(DE3)
4.) Mourez, M. et al. Mapping
k ,mk ) gyrA96 relA1 thi mcrA Δ(lac-proAB) ΔaraB:: ΔR, cat F’[traD36 proAB+ lacIq dominant-negative mutations of
anthrax protective antigen by
Comments: Includes chromosomally encoded bacteriophage lambda R gene and lambda DE DNA to express T7 RNA
scanning mutagenesis, PNAS, Nov
Polymerase. Partly restriction-deficient; good strain for cloning repetitive DNA(recA- ). Suppresses many amber mutations when glutamine is available but not the S100 or S7 mutation of λ, e.g., λgt11. Can be used for M13 cloning/sequencing and blue/white screening.
C600
F- [e14-(McrA-) or e14+ (mcrA+)] thr- 1leuB6 thi- 1 lacY1 supE44 rfbD1 fhuA21; the original C600 is EcoK r+m+ McrBC+ (2,3)

DH5a
F- φ80dlacZΔM15 Δ(lacZYA-argF)U169 deoR, recA1 endA1 hsdR17(r -
Comments: Insert stability due to Reca1 mutation. Can be used for blue/white screening, accepts large plasmids due to
deoR mutation. High plasmid yield due to endA1 mutation.
HB101
F-Δ(gpt-proA)62 leuB6 supE44 ara-14 galkK2 lacY1 Δ(mcrC-mrr) rpsL20 (Strr) xyl-5 mtl-1 recA13 (4)

TG1
F’traD36 lacIq Δ(lacZ) M15 proA+B+ /supE Δ(hsdM-mcrB)5 (r -

Ordering Information

Description
For general cloning, blue-white selection, plasmid isolation. Healthy strain w/ transformation efficiency > 108. For general cloning, blue-white selection, plasmid isolation. Slow growth w/ certain plasmids not stable. Transformation efficiency > 108. For general cloning, plasmid isolation. Transformation For general cloning, plasmid isolation. Transformation For general cloning, blue-white selection, plasmid isolation. JM109 w/ chromosomally inserted λ lysozyme gene that is Autolysis
inducible by arabinose. Z-Competent™ version is chemical y competent with high transformation efficiency. XJa(DE3)
JM109(DE3) with chromosomally inserted λ lysozyme gene Autolysis
inducible by arabinose. DE3 lysogen encodes chromosomally-encoded T7 polymerase and is a suitable host for T7 driven expression of recombinant proteins (such as with the pET system). Z-Competent™ version are chemically competent with high transformation efficiency. * Available as (10) 100 µl single tube aliquots or (12) 50 µl 8-tube strips.
Toll Free: 1-888-882-9682 • Fax: 1-714-288-9643 • Web: www.zymoresearch.com • E-mail: [email protected] Popular E. coli Related Products from Zymo Research
Z-Competent™ E. coli
Includes all buffers for making up to 20 ml Z-Competent™ E. coli. Transformation Kit
Z-Competent™ E. coli
Includes all buffers for making up to 60 ml Z-Competent™ E. coli. Transformation Buffer
A specially formulated E. coli growth medium used in the preparation of highly M3015-100
ZymoBroth™
competent E. coli for DNA transformation purposes. Can increase the transformation efficiency from 5 to 100-fold (depending on the M3015-500
Premade Ampicillin solution. Ampicillin inhibits bacterial cell wall synthesis. Ampicillin Trihydrate
Commonly used to select for Ampicillin-resistant plasmid bearing strains of bacteria. Effective against both gram (-) and gram (+) bacteria. A1001-25
Premade Chloramphenicol solution. Chloramphenicol inhibits bacterial protein synthesis by binding 50S ribosomal subunit. Commonly used for the Chloramphenicol
amplification of vectors in gram (-) bacteria. Effective against both gram (-) and A1002-25
gram (+) bacteria and some mycobacteria. Premade Kanamycin solution. Kanamycin inhibits bacterial protein synthesis by binding 70S ribosomes resulting in dysfunctional translation of mRNA Kanamycin Sulfate
commonly used to select for cosmid vectors. Effective against both gram (-) A1003-25
Premade Tetracycline solution. Tetracycline inhibits bacterial protein synthesis Tetracycline
by binding the 30S ribosomal subunit. Effective against both gram (-) and gram Hydrochloride USP
A1004-25
Rattler™ Plating
Sterile 5 mm plating beads are convenient and easy to use. No flaming required. Spread cells evenly over the entire culture plate surface. *Bulk quantities are available upon request. Please contact: [email protected] or call 1-888-882-9682 for assistance. Toll Free: 1-888-882-9682 • Fax: 1-714-288-9643 • Web: www.zymoresearch.com • E-mail: [email protected]

Source: http://www.biocenter.hu/pdf/T3003.PDF