Invitrogen Zero Blunt TOPO PCR Cloning Kit, without competent cells
Provides a highly efficient, 5-minute, one-step cloning strategy ( in. TOPO Cloning in. ) for the direct insertion of blunt-ended PCR products amplified with proofreading thermostable polymerases into a plasmid vector
Manufacturer: Invitrogen 450245
Includes the Zero Blunt TOPO vector containing the ccdB gene for positive selection, only permitting growth of plasmid vectors with recombinants. Resistant to Kanamycin (KanR) and Zeocin (ZeoR).
pCRBlunt II-TOPO vector: designed for proofreading polymerases
The pCRBlunt II-TOPO vector is designed to clone blunt-ended PCR products generated by thermostable proofreading polymerases such as Platinum Pfx DNA Polymerase. The pCRBlunt II-TOPO vector contains:
- EcoRI sites flanking the PCR product insertion site for easy excision of inserts
- Kanamycin and Zeocin resistance genes for your choice of selection in E. coli
- SP6 promoter/primer site for in vitro RNA transcription and sequencing
- M13 forward and reverse primer sites for sequencing or PCR screening
pCRBlunt II-TOPO allows for direct selection of recombinants via disruption of the lethal E. coli gene, ccdB. The vector contains the ccdB gene fused to the C-terminus of the LacZα fragment. Ligation of a blunt-end PCR product disrupts expression of the LacZα-ccdB gene fusion, permitting growth of only positive recombinants upon transformation. Cells that contain non-recombinant vector are killed upon plating. Therefore, blue/white screening is not required.
Simplified TOPO-based cloning
Using TOPO cloning technology, there is no need for PCR primers containing specific sequences, post-PCR procedures, vector preparation, or other time-intensive DNA manipulation steps. Just add your PCR reaction straight to the provided topoisomerase-charged vector, incubate 5 minutes, and transform E. coli competent cells.
With up to 95% of clones carrying the desired insert, you can screen fewer clones, saving you time and money. The pCRBlunt II-TOPO vector used in this kit comes with lethal ccdB gene for efficient selection of blunt-ended PCR recombinants products generated by thermostable proofreading polymerases.
The standard in cloning
When it comes to cloning, TOPO cloning technology has been a reliable partner for thousands of scientists for over ten years. Fast, simple-to-use, and efficient, TOPO cloning has been applied to many different vectors for a wide array of applications.
Zero Blunt TOPO PCR Cloning Kit options
The Zero Blunt TOPO PCR Cloning Kit for direct insertion of blunt-ended PCR products into a plasmid vector can be purchased with a variety of competent cells that deliver different advantages depending upon your needs
General cloning: TOP10 cells (Cat. No. K2800-20, K2800-40)
High-efficiency cloning: TOP10 Electrocomp Cells (Cat. No. K2860-01, K2860-40)
General cloning, bacteriophage T1 resistance: DH5α-T1R (Cat. Nos. K2820-20, K2820-40)
Fast growth: Mach1-T1R Chemically Competent E. Coli (Cat. No. K2830-20)
Provide your own: for flexibility and to save money (Cat. No. 450254)
A version of the kit that includes a PureLink Quick Plasmid Miniprep Kit (Cat. No. K2800-02) for use in isolation of clean, sequencing-ready, recombinant plasmid is also available.
- Fast and easy: go from PCR to clones in just three steps and in as little as 5 minutes hands-on time
- Efficient: obtain up to 95% clones with correct insert
- Proven: reliable performance for over a decade with over 4,000 citations
- Simple: no ligase, post-PCR procedures, or PCR primers containing specific sequences are required
- Vector: pCRBlunt II-TOPO vector: contains the ccdB gene, allowing for direct selection of recombinants
- Cloning method: TOPO cloning: Topoisomerase Ibased, 5-minute ligation of blunt-ended PCR products amplified with proofreading thermostable polymerases
- Competent cells: Various options: choose from kits with either general, high-efficiency, bacteriophage T1-resistant, fast-growing competent cells (available separately) or use your own
- Antibiotic resistance: Ampicillin (AmpR), Kanamycin (KanR)
Cloning, PCR Cloning
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Shipping Condition: Dry Ice
|Kit, PCR Cloning|
|Box 1: Linearized and topoisomerase I-activated pCRBlunt II-TOPO vector, Salt solution, dNTPs, Control template, M13 forward and reverse primers, Sterile water|
|Cloning, PCR Cloning|
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For Research Use Only. Not for use in diagnostic procedures.