pKD46

Name: pKD46
SKU: OVT2612
Rating: 4.0 (2 reviews)

For research use only. Not intended for any clinical use.

Cat.No.

OVT2612

Selection

Ampicillin

Source

Escherichia coli

Species

Bacterial

Background

Case Study

Applications

Publications

Q & A

Customer Reviews

The pKD46 plasmid is a reputable vector often used in the Red homologous recombination system, and it is currently the most widely employed. This primary system tool consists of three crucial genes: exo, bet, and gam, all of which are mobilized and controlled by the AraBAD promoter. The plasmid's replicon is a low-copy, temperature-sensitive one, thus making the plasmid's elimination easy and straightforward.

Specially designed for recombineering in E. Coli and other bacteria, pKD46 enables chromosomal deletions of genes with FRT sites. The pKD46’s promoter is the araBAD promoter, known for regulating the metabolism of the three genes, while the replicon originates from the pSC101 ori. The lambda tL3 Terminator serves as its terminator. Furthermore, its prokaryotic resistance is Ampicillin Amp, protecting the plasmid from any antibiotic threats. Induction of the pKD46 is achieved with the aid of Arabian sugar. This substance functions as the activator of the araBAD promoter, subsequently triggering the expression of the three integral genes, exo, bet, and gam. 

Homologous recombination efficiency depends on various parameters such as recA gene expression, GC content, arabinose and PCR concentration. Based on this, the researchers evaluated the expression of recA and gam genes in E. coli for homologous recombination efficiency. In this study, hybrid constructs with different GC content, arabinose concentration, and PCR products were prepared by electroporation into standard E. coli using the Wanner protocol. Similarly, to evaluate the expression of recA and gam genes, the pKD46 plasmid was transformed into E. coli. Real-time PCR for E. coli was performed twice in triplicate, and the mean and standard deviation were calculated. Maximum recombinant colonies obtained with 55% GC content.

The results also showed that as the amount of electroporated PCR products increased, the number of recombinant colonies resistant to chloramphenicol also increased. The results showed that there were significant differences in the expression of recA and gam genes between the standard host and the standard host containing the pKD46 vector. Quantitative PCR results showed that the expression levels of recA and gam genes in vector plasmid pKD46 increased more than a hundred times compared with the standard host.

Figure 1. pKD46 vector and functional exo, bet & gam genes. (MARASHI, SM AMIN, et al., 2016)

Gene Deletion: pKD46 vector, a plasmid that carries the Red system, is commonly used for gene deletion in bacteria, especially in Escherichia coli. It allows for the replacement of a chosen gene by a selectable marker.

Genomic Engineering: pKD46 vector is a versatile tool used to facilitate genomic alterations. Researchers can use it to introduce point mutations, insertions, and deletions.

Lambda Red Recombinase System: pKD46 vector expresses the lambda red recombination system. This system significantly enhances the frequency of recombination, making it a useful tool for genetic research.

Expression of Antibiotic Resistance Genes: pKD46 vector contains antibiotic resistance gene. It can be used to express antibiotic resistance in bacteria, thereby allowing for the selection of successfully transformed organisms.

Study of Gene Function: By using pKD46 vector to delete or mutate a particular gene, researchers can study its function. This is critical in understanding the roles of various genes in bacteria.

Customer Q&As

What is pKD46 vector used for?

A: pKD46 vector is widely used for Red homologous recombination and it contains exo, bet, and gam genes regulated by an arabinose promoter.

What is the function of Exo protein?

A: Exo protein is a nucleic acid exonuclease that degrades DNA from the 5' end to the 3' end, creating 3' sticky ends.

What kind of replication system does pKD46 vector have?

A: pKD46 vector has a low-copy temperature-sensitive replication system, which facilitates plasmid elimination.

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Customer Reviews

Efficient recombination

pKD46 is a fantastic genetic tool for bacterial recombination studies. This temperature-sensitive plasmid allows for efficient recombination without the need for complex procedures.

Germany

08/14/2021

Valuable tool

pKD46's temperature-sensitive feature aids in easy curing of the plasmid from the cell, by simply shifting the temperature to a non-permissive condition.

Germany

03/30/2020

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pKD46

Background

Case Study

Applications

Publications

Q & A

Customer Reviews

What is pKD46 vector used for?

What is the function of Exo protein?

What kind of replication system does pKD46 vector have?

Efficient recombination

Valuable tool

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