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TOPO™ TA Cloning™ Kit, Dual Promoter, with pCR™II-TOPO™ Vector and One Shot™ TOP10F' Chemically Competent E. coli (Invitrogen™)

TOPO® TA Cloning® Kits are designed for cloning PCR products directly from a PCR reaction in just 5 minutes (1). They use a pCR™-TOPO® Vector with covalently bound topoisomerase I for fast cloning and recombinants. pCR™-TOPO® Vectors include:

• 3´-T overhangs for direct ligation of Taq-amplified PCR products
• Choice of T7 (pCR™2.1-TOPO®) or T7 and SP6 (pCR™II-TOPO®) promoters for in vitro RNA transcription and sequencing. Each vector also contains M13 forward and reverse primer sites for sequencing.
EcoR I sites flanking the PCR product insertion site for easy excision of inserts
• Kanamycin and ampicillin resistance genes for your choice of selection in E. coli
• Easy blue/white colony screening for selection of recombinants

TOPO® TA Cloning® Kits are also available in combo kits combined with a PureLink™ Quick Plasmid Miniprep Kit (50 preps) for fast plasmid purification of your TOPO®-cloned inserts for downstream analysis.

Zero Blunt™ TOPO™ PCR Cloning Kit for Sequencing, with One Shot™ TOP10 Chemically Competent E. coli (Invitrogen™)

The Zero Blunt® TOPO® PCR Cloning Kit for Sequencing provides a highly efficient, 5-minute, one-step cloning strategy for the direct insertion of proofreading-polymerase–amplified, blunt-ended PCR products into a plasmid vector for sequencing. Each kit uses the pCR™4Blunt-TOPO® vector (see figure) with specially designed sequencing primer sites that return more insert sequence and less vector sequence from each reaction. These kits include everything necessary to clone and select recombinant vectors containing your PCR fragment of choice.

pCR™4Blunt-TOPO® vectoroptimized for sequencing
We have removed much of the multiple cloning site from the pCR™4Blunt-TOPO® vector to shorten the distance between sequencing primer sites and the insert site to as little as 33 bp. This means sequencing reactions give less vector sequence and more insert sequence. The pCR™4Blunt TOPO® vector has sites for 4 common sequencing primers: M13 forward, M13 reverse, T7, and T3. The kits include an aliquot of each.

pCR™4Blunt-TOPO® clone selection and manipulation
The pCR™4Blunt-TOPO® vector contains both ampicillin and kanamycin resistance markers and a LacZα-ccdB gene fusion for positive selection. The vector’s minimized multiple cloning site still includes flanking EcoRI sites for simplified excision of cloned PCR products and a unique Sse8387I site for generation of nested deletions prior to sequencing. T7 and T3 promoters are also present for in vitro transcription.

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 the provided E. coli competent cells.

Efficient cloning
With up to 95% of clones carrying the desired insert, you can screen fewer clones, which saves you time and money. The pCR™4Blunt-TOPO® vector used in this kit comes with no overhangs for efficient ligation of PCR products created by proofreading, thermostable polymerases that leave blunt-ended PCR products.

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 for Sequencing—kit formats
The Zero Blunt® TOPO® PCR Cloning Kit for Sequencing can be purchased with a variety of competent cells that deliver different advantages depending on what your needs are:

• General cloning: TOP10 cells (Cat. No. K2875-J10, K2875-20, K2875-40)
• High-efficiency cloning: TOP10 Electrocomp™ Cells (Cat. No. K2880-20, K2880-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R (Cat. No. K2895-20)
• Fast growth: Mach1™-T1R Chemically Competent E. Coli (Cat. No. K2835-20)

Related Links

Custom Vector Construction and Cloning Services
Plasmid DNA Purification Kit Selection Guide
PCR Reagents, Instruments, and Supplies

TOPO™ TA Cloning™ Kit for Subcloning, with One Shot™ TOP10 chemically competent E. coli cells (Invitrogen™)

TOPO® TA Cloning® Kits for Subcloning provide a highly efficient, 5-minute, one-step cloning strategy ("TOPO® cloning") for the direct insertion of Taq polymerase-amplified PCR products into a plasmid vector for subcloning. Each kit uses the pCR™ 2.1-TOPO® TA vector with convenient restriction sites for subcloning. The cloning kits are available with a variety of competent cells, or without competent cells, depending upon your needs and budget. Features of the TOPO® TA Cloning® Kits for Subcloning:

Fast and easy—go from PCR to clones in just 3 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

TOPO® TA Cloning® Kits for Subcloning—overview

Vector: pCR™ 2.1-TOPO® TA vector—subcloning vector with 15 convenient restriction sites flanking your insert for easy, directional subcloning

Cloning method: TOPO® TA Cloning® —Topoisomerase I-based, 5-minute ligation of PCR products with 3′-A overhangs (Taq polymerase amplified) to TOPO® vector with T overhangs

Competent cells: various options—choose from kits with either general, high-efficiency, bacteriophage T1-resistant, fast-growing competent cells, or use your own

pCR™ 2.1-TOPO® TA vector—cloning and subcloning simplicity
The pCR™ 2.1-TOPO® TA vector is linearized with 3'-thymidine (T) overhangs for direct ligation of Taq polymerase-amplified PCR products (TA cloning®) and is "activated" with covalently bound Topoisomerase I. There is no need to add ligase and cloning is complete in 5 minutes. EcoRI sites flanking the PCR product insertion site allow for easy excision of inserts or use any combination of 15 convenient restriction sites flanking your PCR insert for easy, directional subcloning.

pCR™ 2.1-TOPO® TA clone selection and manipulation
The pCR™ 2.1-TOPO® TA vector contains both ampicillin and kanamycin resistance markers and the LacZα gene for blue/white screening.

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.

Efficient cloning
With up to 95% of clones carrying the desired insert, you can screen fewer clones, saving time and money. The pCR™ 2.1-TOPO® TA vector used in this kit comes with 3'-T overhangs for efficient ligation of Taq polymerase-amplified PCR products, which contain 3'-A overhangs.

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.

TOPO® TA Cloning® Kits for Subcloning—kit options
The TOPO® TA Cloning® Kit for Sequencing can be purchased with a variety of competent cells that deliver different advantages depending upon your needs:

• General cloning: TOP10 cells (Cat. No. K4500-01, K4500-40)
• High-efficiency cloning: TOP10 Electrocomp™ Cells (Cat. No. K4560-01, K4560-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R cells (Cat. No. K4520-01, K4520-40)
• Fast growth: Mach1™ -T1R Chemically Competent E. Coli (Cat. No. K4510-20)
• Provide your own: for flexibility and to save money (Cat. No. 450641)

We also offer two versions of the kit that include a PureLink™ Quick Plasmid Miniprep Kit (Cat. No. K4500-02 and K4510-02) for use in isolation of clean, sequencing-ready, recombinant plasmid.

TOPO™ TA Cloning™ Kit, Dual Promoter, with pCR™II-TOPO™ Vector and One Shot™ TOP10 Electrocomp™ E. coli (Invitrogen™)

TOPO® TA Cloning® Kits are designed for cloning PCR products directly from a PCR reaction in just 5 minutes (1). They use a pCR™-TOPO® Vector with covalently bound topoisomerase I for fast cloning and recombinants. pCR™-TOPO® Vectors include:

• 3´-T overhangs for direct ligation of Taq-amplified PCR products
• Choice of T7 (pCR™2.1-TOPO®) or T7 and SP6 (pCR™II-TOPO®) promoters for in vitro RNA transcription and sequencing. Each vector also contains M13 forward and reverse primer sites for sequencing.
EcoR I sites flanking the PCR product insertion site for easy excision of inserts
• Kanamycin and ampicillin resistance genes for your choice of selection in E. coli
• Easy blue/white colony screening for selection of recombinants

TOPO® TA Cloning® Kits are also available in combo kits combined with a PureLink™ Quick Plasmid Miniprep Kit (50 preps) for fast plasmid purification of your TOPO®-cloned inserts for downstream analysis.

Zero Blunt™ TOPO™ PCR Cloning Kit for Sequencing, with One Shot™ TOP10 Electrocomp™ E. coli (Invitrogen™)

The Zero Blunt® TOPO® PCR Cloning Kits for Sequencing provide a highly efficient, 5 minute, one-step cloning strategy ("TOPO® Cloning") for the direct insertion of proofreading-polymerase–amplified blunt-end PCR products into a plasmid vector for sequencing. Each kit uses the pCR™4Blunt-TOPO® vector with specially designed sequencing primer sites that return more insert sequence and less vector sequence from each reaction. These kits include everything necessary to clone and select recombinant vectors containing your PCR fragment of choice.

Get More Sequence—Allows for more insert sequence and less vector sequence when using standard sequencing primers

Fast and Easy—Go from PCR-to-clones in just 3 steps and in as little as 5 minutes hands-on time
Efficient—Achieve up to 95% clones with correct insert
Proven—Reliable performance for over a decade with over 4000 citations

Zero Blunt® TOPO® Cloning Kits for Sequencing Kit Overview
VECTOR: pCR™4Blunt-TOPO® Vector—Optimized cloning vector for improved sequencing results
CLONING METHOD: TOPO® Cloning —Topoisomerase I based 5 minute ligation of proofreading-polymerase-amplified PCR products with blunt ends to the vector
COMPETENT CELLS: Various Options —Choose from kits with either general, high-efficiency, bacteriophage T1-resistant, or fast-growing competent cells

pCR™4Blunt-TOPO® VectorOptimized for Sequencing
We have removed much of the multiple cloning site from the pCR™4Blunt-TOPO® vector to shorten the distance between sequencing primer sites and the insert site to as little as 33 bp. This means sequencing reactions give less vector sequence and more insert sequence. The pCR™4Blunt TOPO® vector has sites for 4 common sequencing primers: M13 forward, M13 reverse, T7, and T3. The kits include an aliquot of each.

pCR™4Blunt-TOPO® Clone Selection and Manipulation
The pCR™4Blunt-TOPO® vector contains both ampicillin and kanamycin resistance markers and a LacZα-ccdB gene fusion for positive selection. The vector’s minimized multiple cloning site still includes flanking EcoRI sites for simplified excision of cloned PCR products and a unique Sse8387I site for generation of nested deletions prior to sequencing. T7 and T3 promoters are also present for in vitro transcription.

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 the provided E. coli competent cells.

Efficient Cloning
With up to 95% of clones carrying the desired insert, you can screen fewer clones to help you save time and money. The pCR™4Blunt-TOPO® vector used in this kit comes with no overhangs for efficient ligation of PCR products created by proofreading, thermostable polymerases that leave blunt-ended PCR products.

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 Kits for Sequencing Kit Options
The Zero Blunt® TOPO® PCR Cloning Kit for Sequencing can be purchased with a variety of competent cells that deliver different advantages depending on what your needs are:

• General cloning: TOP10 (Cat. No. K2875-J10, K2875-20, K2875-40)
• High efficiency cloning: TOP10 Electrocomp™ Cells (Cat. No. K2880-20, K2880-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R (Cat. No. K2895-20)
• Fast growth: Mach1™-T1R Chemically Competent E. Coli (Cat. No. K2835-20)

TOPO® products are For Research Use Only. Not intended for any animal or human therapeutic of diagnostic use.

Related Links
Custom Vector Construction and Cloning Services
Plasmid DNA Purification Kit Selection Guide
PCR Reagents, Instruments, and Supplies

TOPO™ TA Cloning™ Kit for Subcloning, with TOP10 E. coli (Supply Center Packaging) (Invitrogen™)

TOPO® TA Cloning® Kits for Subcloning provide a highly efficient, 5-minute, one-step cloning strategy ("TOPO® cloning") for the direct insertion of Taq polymerase–amplified PCR products into a plasmid vector for subcloning. Each kit uses the pCR™ 2.1-TOPO® TA vector with convenient restriction sites for subcloning. The cloning kits are available with a variety of competent cells, or without competent cells, depending upon your needs and budget. This product comes in Supply Center packaging, which features an additional outer box that holds both the TOPO® vector box and the competent cells box. This special packaging helps ensure that visitors to a Supply Center obtain the complete kit.

Features of the TOPO® TA Cloning® Kits for Subcloning:

Fast and easy—go from PCR to clones in just 3 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

TOPO® TA Cloning® Kits for Subcloning—overview

Vector: pCR™ 2.1-TOPO® TA vector—subcloning vector with 15 convenient restriction sites flanking your insert for easy, directional subcloning
Cloning method: TOPO® TA Cloning® —Topoisomerase I–based, 5-minute ligation of PCR products with 3′-A overhangs (Taq polymerase amplified) to TOPO® vector with T overhangs
Competent cells: various options—choose from kits with either general, high-efficiency, bacteriophage T1–resistant, fast-growing competent cells, or use your own

pCR™ 2.1-TOPO® TA vector—cloning and subcloning simplicity
The pCR™ 2.1-TOPO® TA vector is linearized with 3'-thymidine (T) overhangs for direct ligation of Taq polymerase–amplified PCR products (TA cloning®) and is "activated" with covalently bound Topoisomerase I. There is no need to add ligase and cloning is complete in 5 minutes. EcoRI sites flanking the PCR product insertion site allow for easy excision of inserts or use any combination of 15 convenient restriction sites flanking your PCR insert for easy, directional subcloning.

pCR™ 2.1-TOPO® TA clone selection and manipulation
The pCR™ 2.1-TOPO® TA vector contains both ampicillin and kanamycin resistance markers and the LacZα gene for blue/white screening.

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.

Efficient cloning
With up to 95% of clones carrying the desired insert, you can screen less clones, saving time and money. The pCR™ 2.1-TOPO® TA vector used in this kit comes with 3'-T overhangs for efficient ligation of Taq polymerase–amplified PCR products, which contain 3'-A overhangs.

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.

TOPO® TA Cloning® Kits for Subcloning—kit options
The TOPO® TA Cloning® Kit for Sequencing can be purchased with a variety of competent cells that deliver different advantages depending upon your needs:

• General cloning: TOP10 cells (Cat. No. K4500-01, K4500-40)
• High-efficiency cloning: TOP10 Electrocomp™ Cells (Cat. No. K4560-01, K4560-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R cells (Cat. No. K4520-01, K4520-40)
• Fast growth: Mach1™ -T1R Chemically Competent E. Coli (Cat. No. K4510-20)
• Provide your own: for flexibility and to save money (Cat. No. 450641)

We also offer two versions of the kit that include a PureLink™ Quick Plasmid Miniprep Kit (Cat. No. K4500-02 and K4510-02) for use in isolation of clean, sequencing-ready, recombinant plasmid.

TA Cloning™ Kit, Dual Promoter, with pCR™II Vector and One Shot™ INVαF' Chemically Competent E. coli (Invitrogen™)

The TA Cloning® Kit Dual Promoter with pCR™II vector provides a quick, one-step cloning strategy for directly inserting a Taq-amplified PCR product into a plasmid vector. The T7 and Sp6 promoters of the pCR™II vector allow in vitro transcription of the insert to produce sense or anti-sense products. The TA Cloning Kit Dual Promoter uses the pCR™II cloning vector and ExpressLink™ T4 DNA Ligase to generate a ligation product in a fifteen-minute, room-temperature ligation step. Reactions typically yield >80% recombinants containing inserts.

Features of the TA Cloning® Kit Dual Promoter:
Fast & convenient—15-minute, room-temperature ligation
Efficient—blue/white screening and >80% clones with correct insert
Flexible—choice of kanamycin or ampicillin resistance for flexible antibiotic selection
Hassle-free—eliminates any enzymatic modifications of the PCR product
Streamlined—does not require the use of PCR primers that contain restriction sites

The pCR™II vector provides:
• 3'-T overhangs for direct ligation of Taq-amplified PCR products
• T7 and Sp6 promoters for in vitro RNA transcription and sequencing
• Versatile polylinker with flanking EcoR I sites for easy excision of inserts
• M13 forward and reverse primer sites for sequencing

How TA Cloning® Works
Taq polymerase has a non-template-dependent activity that adds a single deoxyadenosine (A) to the 3' ends of PCR products. The linearized vector supplied in this kit has single 3' deoxythymidine (T) residues. This allows PCR inserts to ligate efficiently with the vector.

Kit Configurations
The TA Cloning® Kit is offered in a variety of configurations: with One Shot® INVF' Chemically Competent E. coli (K2050-01 and K2050-40), One Shot® TOP10F' Chemically Competent E. coli (K2060-01 and K2060-40), and without competent cells (K2750-20 and K2750-40) in 20- and 40-reaction kit sizes.

TOPO™ TA Cloning™ Kit for Sequencing, without competent cells (Invitrogen™)

TOPO® TA Cloning® kits for sequencing provide a highly efficient, 5-minute cloning reaction for the direct insertion of Taq polymerase-amplified PCR products into a plasmid vector for sequencing. Each kit uses the pCR™4-TOPO® TA vector with specially designed sequencing primer sites that return more insert sequence and less vector sequence from each reaction, and are available with a variety of competent cells, or no competent cells, depending on your needs and budget. These kits include the tools necessary to clone and select recombinant vectors containing your PCR fragment of choice.

• Get more sequence—allows for more insert sequence and less vector sequence when using standard sequencing primers
• Fast and easy—go from PCR to clones in just 3 steps, including a 5-minute cloning reaction
• Efficient—obtain up to 95% clones with correct insert
• Proven—enabling reliable performance for over a decade with over 20,000 citations

pCR™4-TOPO® TA Vector—Optimized for Sequencing
The pCR™4-TOPO® TA vector has a minimized multiple cloning site and, thus, a shorter distance between the sequencing primer sites and the insert site (as little as 33 bp). So sequencing reactions produce less vector sequence and more insert sequence. The pCR™4-TOPO® TA vector has sites for 4 common sequencing primers: M13 forward, M13 reverse, T7, and T3. The kits include an aliquot of each.

pCR™4-TOPO® TA Clone Selection and Manipulation
The pCR™4-TOPO® TA vector contains both ampicillin and kanamycin resistance markers and a LacZα-ccdB gene fusion for positive selection and blue/white screening. The vector’s minimized multiple cloning site still includes flanking EcoRI sites for simplified excision of cloned PCR products and a unique Sse8387I site for generation of nested deletions prior to sequencing. T7 and T3 promoters are also present for in vitro transcription.

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.

Efficient Cloning
With up to 95% of clones carrying the desired insert, you can screen less clones and save time and money. The pCR™4-TOPO® TA vector used in this kit comes with 3´T overhangs for efficient ligation of Taq-amplified PCR products, which contain 3´A overhangs.

The Most Widely Used Cloning Kit
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.

Kit Options: TOPO® TA Cloning® Kits for Sequencing
TOPO® TA Cloning® kits for sequencing can be purchased with a variety of competent cells that deliver different advantages depending upon your needs:
• General cloning: TOP10 (Cat. Nos. K4575-J10, K4575-01, K4575-40)
• High-efficiency cloning: TOP10 Electrocomp™ cells (Cat. Nos. K4580-01, K4580-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R (Cat. Nos. K4595-01, K4595-40)
• Fast growth: Mach1™-T1R chemically competent E. coli (Cat. No. K4530-20)
• Provide your own cells (Cat. Nos. 450071, 450030)

We also offer a version of the kit that includes a PureLink® Quick Plasmid Miniprep Kit (Cat. No. K4575-02) for use in isolation of clean, sequencing-ready, recombinant plasmid.

TOPO™ TA, Dual TOP10 (Supply Center Packaging) (Invitrogen™)

The TOPO® TA Cloning® Dual Promoter Kit is for fast, efficient cloning and subsequent in vitro transcription/translation. This kit includes the pCR™II-TOPO® TA vector (see figure). By eliminating time-consuming and tedious restriction site cloning, TOPO® cloning is the most reliable method, taking only 5 minutes using a 3-step protocol, and yielding up to 95% recombinants. Convenient features of the TOPO® TA Cloning® Dual Promoter Kit include:

• 3´-T overhangs for direct ligation of Taq-amplified PCR products
• T7 and SP6 promoters for efficient in vitro transcription
• M13 forward and reverse primer sites for sequencing
• 16 convenient restriction sites, including EcoRI, flanking your insert for subsequent excision or subcloning
• Kanamycin and ampicillin resistance for your choice of selection in E. coli
• Easy blue/white colony screening for selection of recombinants
• Includes OneShot® TOP10 Competent cells for added convenience and highest cloning efficiencies

This product comes in Supply Center packaging, which features an additional outer box that holds both the TOPO® vector box and the competent cells box. This special packaging helps ensure that visitors to a Supply Center obtain the complete kit.

TOPO™ TA Cloning™ Kit, with pCR™2.1-TOPO™, One Shot TOP10 Chemically Competent E. coli, and PureLink™ Quick Plasmid Miniprep Kit (Invitrogen™)

TOPO® TA Cloning® Kits are designed for cloning PCR products directly from a PCR reaction in just 5 minutes (1). They use a pCR™-TOPO® Vector with covalently bound topoisomerase I for fast cloning and recombinants. pCR™-TOPO® Vectors include:
• 3´-T overhangs for direct ligation of Taq-amplified PCR products
• Choice of T7 (pCR™2.1-TOPO®) or T7 and SP6 (pCR™II-TOPO®) promoters for in vitro RNA transcription and sequencing. Each vector also contains M13 forward and reverse primer sites for sequencing.
EcoR I sites flanking the PCR product insertion site for easy excision of inserts
• Kanamycin and ampicillin resistance genes for your choice of selection in E. coli
• Easy blue/white colony screening for selection of recombinants

TOPO® TA Cloning® Kits are available in combo kits combined with a PureLink™ Quick Plasmid Miniprep Kit (50 preps) for fast plasmid purification of your TOPO®-cloned inserts for downstream analysis.

Zero Blunt™ PCR Cloning Kit (Invitrogen™)

The Zero Blunt® PCR Cloning Kit offers an easy method for high-efficiency (>80%) cloning of blunt-end PCR products amplified with proof-reading, thermostable DNA polymerases. The Zero Blunt® PCR Cloning Kit uses the multipurpose cloning vector pCR™-Blunt and ExpressLink™ T4 DNA Ligase to generate a ligation product in a five-minute, room-temperature ligation step.

Features of the Zero Blunt® Cloning® Kit with pCR™-Blunt vector:
Fast & convenient—5-minute, room-temperature ligation
EfficientccdB gene for positive selection results in >80% clones with correct insert
Flexible—choice of kanamycin or Zeocin™ resistance for flexible antibiotic selection

The pCR™-Blunt vector provides:
EcoR I sites flanking the PCR product insertion site for excision of inserts
• T7 promoter/primer site for in vitro RNA transcription and sequencing
• M13 forward and reverse primer sites for sequencing or PCR screening

How Zero Blunt® PCR Cloning Works
The Zero Blunt® PCR Cloning Kit is designed to clone blunt PCR fragments (or any blunt DNA fragment) with a low background of non-recombinants. The pCR™-Blunt vector contains the lethal E. coliccdB gene fused to the C-terminus of LacZα (Bernard et al., 1994). Ligation of a blunt PCR fragment disrupts expression of the lacZα-ccdB gene fusion permitting growth of only positive recombinants upon transformation. Cells that contain non-recombinant vector are killed when the transformation mixture is plated.

Kit Configurations
The Zero Blunt® PCR Cloning Kit is offered in a variety of configurations: with One Shot® TOPO10 Chemically Competent E. coli (K2700-20 and K2700-40) and without competent cells (K2750-20 and K2750-40) in 20- and 40- reaction kit sizes.

Zero Blunt™ TOPO™ PCR Cloning Kit, with One Shot™ MAX Efficiency DH5α-T1R E. coli (Invitrogen™)

Zero Blunt® TOPO® PCR Cloning Kits offers the fastest and easiest method for high-efficiency (cloning of blunt-end PCR products amplified with proofreading thermostable polymerases. The kits include linearized and topoisomerase I-activated pCR™-Blunt II-TOPO® Vector for 5-minute benchtop ligations without ligase (1). The pCR™-Blunt II-TOPO® Vector also features:

ccdB gene for positive selection (2)
EcoR I 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

Selected Zero Blunt® TOPO® PCR Cloning Kits are available in combo kits combined with a PureLink™ Quick Plasmid Miniprep Kit (50 preps) for fast plasmid purification of your TOPO®-cloned inserts for downstream analysis.

TOPO™ TA Cloning™ Kit for Sequencing, with One Shot™ Mach1™ T1 Phage-Resistant Chemically Competent E. coli (Invitrogen™)

The TOPO® TA Cloning® Kits for Sequencing provide a highly efficient, 5 minute, one-step cloning strategy ("TOPO® Cloning") for the direct insertion of Taq polymerase-amplified PCR products into a plasmid vector for sequencing. Each kit uses the pCR™4-TOPO® TA vector with specially designed sequencing primer sites that return more insert sequence and less vector sequence from each reaction. These kits include everything necessary to clone and select recombinant vectors containing your PCR fragment of choice.

Get More Sequence—Allows for more insert sequence and less vector seuquence when using standard sequencing primers
Fast and Easy—Go from PCR-to-clones in just 3 steps and in as little as 5 minutes hands-on time
Efficient—Achieve up to 95% clones with correct insert
Proven—Reliable performance for over a decade with over 4000 citations

TOPO® TA Cloning® Kits for Sequencing Kit Overview
VECTOR: pCR™4-TOPO® TA Vector—Optimized cloning vector for improved sequencing results
CLONING METHOD: TOPO® TA Cloning —Topoisomerase I based 5 minute ligation of PCR products with 3´A overhangs (Taq-amplified) to the vector
COMPETENT CELLS: Various Options —Choose from kits with either general, high-efficiency, bacteriophage T1-resistant, or fast-growing competent cells

pCR™4-TOPO® TA VectorOptimized for Sequencing
We have removed much of the multiple cloning site from the pCR™4-TOPO® TA vector to shorten the distance between sequencing primer sites and the insert site to as little as 33 bp. This means sequencing reactions give less vector sequence and more insert sequence. The pCR™4-TOPO® TA vector has sites for 4 common sequencing primers: M13 forward, M13 reverse, T7, and T3. The kits include an aliquot of each.

pCR™4-TOPO® TA Clone Selection and Manipulation
The pCR™4-TOPO® TA vector contains both ampicillin and kanamycin resistance markers and a LacZα-ccdB gene fusion for positive selection and blue/white screening. The vector’s minimized multiple cloning site still includes flanking EcoRI sites for simplified excision of cloned PCR products and a unique Sse8387I site for generation of nested deletions prior to sequencing. T7 and T3 promoters are also present for in vitro transcription.

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 with the providedE. coli competent cells.

Efficient Cloning
With up to 95% of clones carrying the desired insert, you can screen less clones to help you save time and money. The pCR™4-TOPO® TA vector used in this kit comes with 3´T overhangs for efficient ligation of Taq-amplified PCR products with 3´A overhangs.

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.

TOPO® TA Cloning® Kits for Sequencing Kit Options
The TOPO® TA Cloning® Kit for Sequencing can be purchased with a variety of competent cells that deliver different advantages depending on what your needs are:

• General cloning: TOP10 (Cat. No. K4575-J10, K4575-01, K4575-40)
• High efficiency cloning: TOP10 Electrocomp™ Cells (Cat. No. K4580-01, K4580-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R (Cat. No. K4595-01, K4595-40)
• Fast growth: Mach1™-T1R Chemically Competent E. Coli (Cat. No. K4530-20)

We also offer a version of the kit that includes a PureLink™ Quick Plasmid Miniprep Kit (Cat. No. K4575-02) for use in isolation of clean, sequencing-ready, recombinant plasmid.

TOPO® products are For Research Use Only. Not intended for any animal or human therapeutic of diagnostic use.

Related Links
Custom Vector Construction and Cloning Services
Plasmid DNA Purification Kit Selection Guide
PCR Reagents, Instruments, and Supplies

TOPO™ TA Cloning™ Kit for Subcloning, with One Shot™ TOP10 Electrocomp™ E. coli (Invitrogen™)

TOPO® TA Cloning® Kits for Subcloning provide a highly efficient, 5-minute, one-step cloning strategy ("TOPO® cloning") for the direct insertion of Taq polymerase–amplified PCR products into a plasmid vector for subcloning. Each kit uses the pCR™ 2.1-TOPO® TA vector with convenient restriction sites for subcloning. The cloning kits are available with a variety of competent cells, or without competent cells, depending upon your needs and budget. Features of the TOPO® TA Cloning® Kits for Subcloning:

Fast and easy—go from PCR to clones in just 3 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

TOPO® TA Cloning® Kits for Subcloning—overview

Vector: pCR™ 2.1-TOPO® TA vector—subcloning vector with 15 convenient restriction sites flanking your insert for easy, directional subcloning

Cloning method: TOPO® TA Cloning® —Topoisomerase I–based, 5-minute ligation of PCR products with 3′-A overhangs (Taq polymerase amplified) to TOPO® vector with T overhangs

Competent cells: various options—choose from kits with either general, high-efficiency, bacteriophage T1–resistant, fast-growing competent cells, or use your own

pCR™ 2.1-TOPO® TA vector—cloning and subcloning simplicity
The pCR™ 2.1-TOPO® TA vector is linearized with 3'-thymidine (T) overhangs for direct ligation of Taq polymerase–amplified PCR products (TA cloning®) and is "activated" with covalently bound Topoisomerase I. There is no need to add ligase and cloning is complete in 5 minutes. EcoRI sites flanking the PCR product insertion site allow for easy excision of inserts or use any combination of 15 convenient restriction sites flanking your PCR insert for easy, directional subcloning.

pCR™ 2.1-TOPO® TA clone selection and manipulation
The pCR™ 2.1-TOPO® TA vector contains both ampicillin and kanamycin resistance markers and the LacZα gene for blue/white screening.

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.

Efficient cloning
With up to 95% of clones carrying the desired insert, you can screen less clones, saving time and money. The pCR™ 2.1-TOPO® TA vector used in this kit comes with 3'-T overhangs for efficient ligation of Taq polymerase–amplified PCR products, which contain 3'-A overhangs.

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.

TOPO® TA Cloning® Kits for Subcloning—kit options
The TOPO® TA Cloning® Kit for Sequencing can be purchased with a variety of competent cells that deliver different advantages depending upon your needs:

• General cloning: TOP10 cells (Cat. No. K4500-01, K4500-40)
• High-efficiency cloning: TOP10 Electrocomp™ Cells (Cat. No. K4560-01, K4560-40)
• General cloning, bacteriophage T1 resistance: DH5α-T1R cells (Cat. No. K4520-01, K4520-40)
• Fast growth: Mach1™ -T1R Chemically Competent E. Coli (Cat. No. K4510-20)
• Provide your own: for flexibility and to save money (Cat. No. 450641)

We also offer two versions of the kit that include a PureLink™ Quick Plasmid Miniprep Kit (Cat. No. K4500-02 and K4510-02) for use in isolation of clean, sequencing-ready, recombinant plasmid.

TA Cloning™ Kit, Dual Promoter, with pCR™II Vector and One Shot™ TOP10F' Chemically Competent E. coli (Invitrogen™)

The TA Cloning® Kit Dual Promoter with pCR™II vector provides a quick, one-step cloning strategy for directly inserting a Taq-amplified PCR product into a plasmid vector. The T7 and Sp6 promoters of the pCR™II vector allow in vitro transcription of the insert to produce sense or anti-sense products. The TA Cloning Kit Dual Promoter uses the pCR™II cloning vector and ExpressLink™ T4 DNA Ligase to generate a ligation product in a fifteen-minute, room-temperature ligation step. Reactions typically yield >80% recombinants containing inserts.

Features of the TA Cloning® Kit Dual Promoter:
Fast & convenient—15-minute, room-temperature ligation
Efficient—blue/white screening and >80% clones with correct insert
Flexible—choice of kanamycin or ampicillin resistance for flexible antibiotic selection
Hassle-free—eliminates any enzymatic modifications of the PCR product
Streamlined—does not require the use of PCR primers that contain restriction sites

The pCR™II vector provides:
• 3'-T overhangs for direct ligation of Taq-amplified PCR products
• T7 and Sp6 promoters for in vitro RNA transcription and sequencing
• Versatile polylinker with flanking EcoR I sites for easy excision of inserts
• M13 forward and reverse primer sites for sequencing

How TA Cloning® Works
Taq polymerase has a non-template-dependent activity that adds a single deoxyadenosine (A) to the 3' ends of PCR products. The linearized vector supplied in this kit has single 3' deoxythymidine (T) residues. This allows PCR inserts to ligate efficiently with the vector.

Kit Configurations
The TA Cloning® Kit is offered in a variety of configurations: with One Shot® INVF' Chemically Competent E. coli (K2050-01 and K2050-40), One Shot® TOP10F' Chemically Competent E. coli (K2060-01 and K2060-40), and without competent cells (K2750-20 and K2750-40) in 20- and 40-reaction kit sizes.