For direct cloning of PCR products generated by Taq DNA polymerases
  • Just 40 minutes from PCR product to plated cells
  • Ready-to-use Ligation Master Mix
  • High-specificity UA hybridization for efficient cloning
  • Immediate plating of transformed competent cells

The QIAGEN PCR Cloning Kit provides ready-to-use ligation reactions, which contain linearized cloning vectors that carry U overhangs at each 3' end, allowing PCR products containing 3'-end A overhangs to be directly ligated and cloned with high efficiency and speed.

Quantity Product Cat. no. Price
 
 
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Quantity Product Cat. no. Price Sum
 
QIAGEN PCR Cloning Kit (10)
For 10 reactions: 2x Ligation Master Mix (50 µl), pDrive Cloning Vector (0.5 µg), Distilled water (1.7 ml)
231122
$97.60
$0.00
 
QIAGEN PCR Cloning Kit (40)
For 40 reactions: 2x Ligation Master Mix (200 µl), pDrive Cloning Vector (2.0 µg), Distilled water (1.7 ml)
231124
$343.00
$0.00
The QIAGEN PCR Cloning Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Highly specific cloning with a shorter ligation time of 30 min.|Highly specific cloning with a ligation time of 4 h.|The QIAGEN PCR Cloning Kit procedure.|pDrive Cloning Vector.|
The effect of ligation time on cloning efficiency was compared for the QIAGEN PCR Cloningplus Kit and a TA-based cloning kit (Supplier I) using PCR products of different length (0.5 kb, 1 kb, and 3 kb). The recommended protocol for each kit was followed. Colony numbers were converted to relative percentages, with the QIAGEN PCR Cloningplus Kit procedure set at 100% for each comparison. The ligation was performed for 30 min (QIAGEN PCR Cloningplus Kit recommendation).|The effect of ligation time on cloning efficiency was compared for the QIAGEN PCR Cloningplus Kit and a TA-based cloning kit (Supplier I) using PCR products of different length (0.5 kb, 1 kb, and 3 kb). The recommended protocol for each kit was followed. Colony numbers were converted to relative percentages, with the QIAGEN PCR Cloningplus Kit procedure set at 100% for each comparison. The ligation was performed for 4 hours (TA-based cloning recommendation).|Simply mix the PCR product directly with pDrive Cloning Vector and Ligation Master Mix, incubate, and then add the ligation reaction to competent cells for transformation.|The pDrive Cloning Vector has a number of useful features designed to facilitate analysis of cloned PCR products. These include a large number of unique restriction enzyme recognition sites, universal sequencing primer sites, and promoters for in vitro transcription. In addition, the vector allows both ampicillin and kanamycin selection, as well as blue/white screening of recombinant colonies.|
Performance

The QIAGEN PCR Cloning Kit combines the latest ligation technology with a unique combination of time-saving features for fast, easy, and highly efficient cloning of PCR products generated using Taq and other nonproofreading DNA polymerases. The QIAGEN PCR Cloning Kit outperformed kits tested from other suppliers, ensuring successful results. Cloning into the pDrive Cloning Vector is much faster compared to TA-based cloning vectors (see figures "Highly specific cloning with a shorter ligation time of 30 min" and "Highly specific cloning with a ligation time of 4 h", and table).

Time from PCR product to plated cells for different cloning methods
QIAGEN PCR Cloning Kit Topoisomerase-mediated cloning kit TA-based cloning kitConventional ligase cloning
40 min ≥70 min ≥5.5 h ≥7.5 h

Principle

The pDrive Cloning Vector (see figure "pDrive Cloning Vector") provides highly efficient cloning of PCR products through UA hybridization. The vector is supplied in a linear form with a U overhang at each 3' end, which hybridizes with high specificity to the A overhang of PCR products generated by Taq and other nonproofreading DNA polymerases. The pDrive Cloning Vector (see figure "pDrive Cloning Vector") has a number of useful features designed to facilitate analysis of cloned PCR products. These include a large number of unique restriction enzyme recognition sites, universal sequencing primer sites, and promoters for in vitro transcription. In addition, the vector allows both ampicillin and kanamycin selection, as well as blue/white screening of recombinant colonies.

PCR products are efficiently cloned into the pDrive Cloning Vector in less time than is required for TA-based cloning vectors (see figures "Highly specific cloning with a shorter ligation time of 30 min" and "Highly specific cloning with a ligation time of 4 h"). Furthermore, as T is the most likely base to hybridize to noncomplementary bases (i.e., G, C, and T), vectors with a T overhang are more likely to self-anneal or to clone primers or annealed primers, leading to an increased number of false-positive colonies. In contrast, the higher cloning efficiency of the pDrive Cloning Vector indicates that U has a lower tolerance for nonspecific base pairing.

The Ligation Master Mix, supplied in a convenient premixed format, is specifically designed to provide optimal hybridization conditions for efficient cloning.

Components included with the QIAGEN PCR Cloning Kit
ComponentIncluded Concentration
pDrive Cloning Vector 50 ng/µl
Ligation Master Mix 2x solution
Distilled water
QIAGEN EZ Competent Cells
SOC medium

 

Procedure

Simply mix the PCR product directly with pDrive Cloning Vector and Ligation Master Mix, incubate, and then add the ligation reaction to competent cells for transformation.

The QIAGEN PCR Cloning Kit procedure is much faster than topoisomerase-mediated, TA-based, and conventional sticky- and blunt-end cloning methods. Ligation takes 30 minutes and transformation and plating using QIAGEN EZ Competent Cells takes only 10 minutes, making the complete procedure — from PCR product to plated cells — just 40 minutes.

 

 

Applications
The QIAGEN PCR Cloning Kit is suitable for cloning of any PCR product that has a single A overhang at each 3' end. PCR products generated using Taq and other nonproofreading DNA polymerases can be directly cloned without any preparation. Kits offering proofreading DNA polymerases, such as the HotStar HiFidelity Polymerase Kit and the QIAGEN LongRange PCR Kit, generate PCR products with A overhangs, and are also highly suited for direct use with the QIAGEN PCR Cloning Kit.
Feature
Specifications
Applications Cloning of A overhang PCR products
Competent cells Not included
Overhang U overhang
Reaction type UA hybridization
Vector included pDrive Cloning Vector

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