One Shot™ TOP10 Chemically Competent E. coli
One Shot&trade; TOP10 Chemically Competent <i>E. coli</i>
Invitrogen™

One Shot™ TOP10 Chemically Competent E. coli

One Shot TOP10 Chemically Competent E. coli are ideal for high-efficiency cloning and plasmid DNA propagation and are provided atRead more
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Catalog NumberQuantity
C40401011 x 50 μL
C40400321 x 50 μL
C40400642 x 50 μL
Catalog number C404010
Price (USD)
310.00
Each
Add to cart
Quantity:
11 x 50 μL
Price (USD)
310.00
Each
Add to cart

One Shot TOP10 Chemically Competent E. coli are ideal for high-efficiency cloning and plasmid DNA propagation and are provided at a transformation efficiency of 1 x 109 cfu/μg plasmid DNA. These cells allow stable replication of high-copy number plasmids and are the same competent cells that come with many of our cloning kits. TOP10 E. coli cells are genetically similar to the DH10B strain and have been reported to be more resilient to stress conditions like osmotic shock and acidic pH stress.

One Shot TOP10 cells:
• Maximize cloning efficiency in a single-tube format
• Provide enhanced genomic DNA cloning capabilities

TOP10 Chemically Competent E. coli cells carry mutations in the methylation-dependent restriction system (mcrA, mcrBC, and mrr) allowing the cloning of both prokaryotic and eukaryotic genomic DNA, as well as efficient plasmid rescue from eukaryotic genomes. Similar to other DH strains, TOP10 has the lacZΔM15 genotype, providing for the option of blue-white screening on plates containing either X-Gal or Bluo-Gal. The inclusion of recA1 and endA1 mutations increase insert stability and improve the quality of plasmid DNA prepared from minipreps.

One Shot TOP10 Chemically Competent E. coli cells offer:
• Transformation efficiencies of >1 x 109 cfu/μg
• hsdR for efficient transformation of unmethylated DNA from PCR amplifications
• mcrA for efficient transformation of methylated DNA from genomic preparations
• lacZΔM15 for blue/white color screening of recombinant clones
• endA1 for cleaner DNA preparations and better results in downstream applications due to elimination of nonspecific digestion by Endonuclease I
• recA1 for reduced occurrence of nonspecific recombination in cloned DNA
• Expression from the lac promoter without IPTG

Easy-to-use One Shot format
TOP10 Chemically Competent E. coli cells are supplied in the convenient, single-reaction One Shot format. The single-tube, single-use format accomodates allows all steps of the transformation protocol, up to plating, to take place in the same tube, helping save time and prevent contamination.

Genotype
F–mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 recA1 araD139 Δ(ara-leu)7697 galU galK λ–rpsL(StrR) endA1 nupG

Find the strain and format that fit your needs 
We offer other DH strains in chemically competent and electrocompetent cell formats.
The TOP10 strain is available in several MultiShot formats for high throughput applications.
Explore bacterial growth media formats.

For Research Use Only. Not for use in diagnostic procedures.
Specifications
Product TypeChemically Competent Cells
Contains F' EpisomeNo
Improves Plasmid QualityYes (endA1)
Cloning Methylated DNAYes (mcrA)
Transformation Efficiency LevelHigh Efficiency (>1 x 109 cfu/μg)
Antibiotic Resistance BacterialYes (Streptomycin)
Cloning Unstable DNANo
Blue/White ScreeningYes (lacZΔM15)
High-throughput CompatibilityLow
PlasmidHigh Copy Plasmid
Preparing Unmethylated DNANo
Reduces RecombinationYes (recA1)
Shipping ConditionDry Ice
T1 Phage - Resistant (tonA)No
SpeciesE. coli (K12)
FormatTube
Product LineOne Shot™
Quantity11 x 50 μL
Unit SizeEach
Contents & Storage
• One Shot TOP10 Chemically Competent E. coli (11 x 50 μL)
Store Competent Cells at –80°C.

• pUC19 DNA (50 μL at 10 pg/μL)
Store pUC19 DNA at –20°C.

• S.O.C. medium (6 mL)
Store S.O.C. Medium at 4°C or room temperature.

Frequently asked questions (FAQs)

What is the difference between TOP10 and TOP10F' cells?

The only difference between TOP10 and TOP10F' cells is that the latter contain the F' episome that carries the tetracycline resistance gene and allows isolation of single-stranded DNA from vectors that have an f1 origin of replication. The F' episome also carries the lacIq repressor for inducible expression from trc, tac, and lac promoters using IPTG. TOP10F' cells require IPTG induction for blue/white screening.

I am trying to clone an insert that is supposedly pretty toxic. I used DH5? and TOP10 cells for the transformation and got no colonies on the plate. Do you have any suggestions for me?

If the insert is potentially toxic to the host cells, here are some suggestions that you can try:

- After transforming TOP10 or DH5? cells, incubate at 25-30°C instead of 37°C. This will slow down the growth and will increase the chances of cloning a potentially toxic insert.
- Try using TOP10F' cells for the transformation, but do not add IPTG to the plates. These cells carry the lacIq repressor that represses expression from the lac promoter and so allows cloning of toxic genes. Keep in mind that in the absence of IPTG, blue-white screening cannot be performed.
- Try using Stbl2 cells for the transformation.

What generation is your ViraPower lentiviral expression system? Can I use it with a 2nd generation lentiviral packaging mix?

Our ViraPower lentiviral expression system is a 3rd generation system with regard to safety features. Our lentiviral expression vectors are derived from wild type HIV, but nearly all the wild type viral proteins (e.g., Vpr, Vpu, Vif, Nef, Tat) have been removed and the HIV envelope is not used. VSV-G (vesicular stomatitis virus G) envelope protein is used instead. Our ViraPower lentiviral expression system can be used with a 2nd generation lentiviral packaging mix. However, our lentiviral packaging mix would not be compatible with a 2nd generation lentiviral expression vector.

What advantages do your Stbl2 cells offer over other cloning strains?

There are other strains available that may function similarly to Stbl2 cells in stabilizing inserts or vectors with repeated DNA sequences. However, one advantage of Stbl2 cells over many similar strains is that they are sensitive to Kanamycin, so you can use Stbl2 to propagate plasmids containing a Kanamycin resistance marker. 

How do you recommend that I prepare my DNA for successful electroporation of E. coli?

For best results, DNA used in electroporation must have a very low ionic strength and a high resistance. A high-salt DNA sample may be purified by either ethanol precipitation or dialysis.

The following suggested protocols are for ligation reactions of 20ul. The volumes may be adjusted to suit the amount being prepared.

Purifying DNA by Precipitation: Add 5 to 10 ug of tRNA to a 20ul ligation reaction. Adjust the solution to 2.5 M in ammonium acetate using a 7.5 M ammonium acetate stock solution. Mix well. Add two volumes of 100 % ethanol. Centrifuge at 12,000 x g for 15 min at 4C. Remove the supernatant with a micropipet. Wash the pellet with 60ul of 70% ethanol. Centrifuge at 12,000 x g for 15 min at room temperature. Remove the supernatant with a micropipet. Air dry the pellet. Resuspend the DNA in 0.5X TE buffer [5 mM Tris-HCl, 0.5 mM EDTA (pH 7.5)] to a concentration of 10 ng/ul of DNA. Use 1 ul per transformation of 20 ul of cell suspension.

Purifying DNA by Microdialysis: Float a Millipore filter, type VS 0.025 um, on a pool of 0.5X TE buffer (or 10% glycerol) in a small plastic container. Place 20ul of the DNA solution as a drop on top of the filter. Incubate at room temperature for several hours. Withdraw the DNA drop from the filter and place it in a polypropylene microcentrifuge tube. Use 1ul of this DNA for each electrotransformation reaction.

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