M13KE Phage Vector is a particularly engineered derivative of M13mp19, specifically designed for expressing peptides as N-terminal pIII fusions in phage display applications. The vector is primarily used for the construction of libraries that are pentavalent, which means that all five copies of pIII in the mature virion carry the fused peptide.
This vector is characterized by the changes it presents as compared to the parent M13mp19. Namely, sites for Acc65I/KpnI and EagI have been introduced, flanking the pIII leader peptidase cleavage site. Additionally, the Acc65I/KpnI site in the multiple cloning site (MCS) has been deleted. The vector plays a crucial role when constructing phage-displayed random peptide libraries, where a synthetic oligonucleotide encoding the random peptide library and a portion of the pIII leader sequence are annealed to an extension primer. Following an extension with DNA polymerase, there is subsequent digestion with Acc65I and EagI. The duplex produced from this process is then inserted into the M13KE Phage Vector that has been digested with the same enzymes.
Viruses are transforming the fields of biosensing and biomedicine due to their multivalency, orthogonal reactivity, and responsiveness to genetic modifications. As the most extensively studied phage model for constructing phage display libraries, M13 phages have received extensive research attention as building blocks or viral scaffolds for a variety of applications including isolation/isolation, sensing/detection, and in vivo imaging. Through genetic engineering and chemical modification, M13 phage can be functionalized into a multifunctional analysis platform, and each functional area can perform its functions without interfering with each other. Its unique filamentous morphology and flexibility also improve analytical performance in terms of target affinity and signal amplification.
Immunoassays are the most dominant form of phage-based assays, in which target-bound M13 acts as a "capture antibody" or "signaling antibody." M13-based capture antibodies exploit the multivalency of the pVIII display system to enhance analyte capture, while M13-based signaling antibodies benefit from the 5:2700 molecular ratio between pIII and pVIII for signal amplification. Wu et al inserted the optimized tetracysteine sequence (TC tag) into the small coat protein coding region of M13KE. The displayed TC tag is displayed on M13 and enters the host cell through infection. TC-tag generates a strong fluorescent signal when combined with fluorescein arsenic helix binder (FlAsH), a membrane-permeable bis-arsenic dye, which facilitates host cell monitoring and viability identification under flow cytometry (Figure 1).
Figure 1. Construction of TC-phage-FlAsH structure for bacterial detection. (Wang R, et al. 2023)
M13KE Phage has an extensive range of applications in the fields of molecular biology and biotechnology. Here are some of the major applications:
Protein Engineering: M13KE phage is manipulated to express altered or novel proteins on its surface. This phage display technology allows libraries of billions of different peptide sequences to be rapidly screened for high-affinity binding to an enormous range of target molecules.
Molecular Cloning: M13KE Phage is consistently used as a cloning vector in molecular biology. Scientists manipulate the phage to carry a small fragment of DNA from another organism. It allows them to produce large quantities of that particular DNA sequence for further analysis or manipulation.
Sequencing Technologies: M13KE Phage has also been used in the development of sequencing technologies. Because of its easy manipulation, it can be used as a clone for whole-genome libraries, from which any desired region of genetic material can be sequenced.
Nanotechnology: The M13KE Phage is also used in the field of nanotechnology. The phage exhibits an inherently superior capability for organizing themselves into highly ordered, resilient, and customizable structures and as a result, it has been used for the construction of nanoscale devices.
Vaccine Development: The surface protein of phage M13KE can be engineered to display a specific antigenic determinant. This means vaccines can be developed that stimulate a strong immune response against that specific determinant.
Mutation Studies: Since M13KE Phage can accommodate foreign DNA and it replicates quickly, it enables researchers to conduct mutation studies efficiently. This helps in understanding various diseases and developing potential remedies.
Customer Q&As
What is M13KE?
A: M13KE is a simple derivative of the M13 bacteriophage, designed for displaying short peptide sequences as fusions to the N-terminus of the pIII protein.
How does M13KE work for displaying peptides?
A: M13KE introduces cloning sites at the 5' end of gene III, allowing short peptides to be fused to pIII. Each virion displays five copies of pIII, all of which are fused to the cloned peptide.
Can I use M13KE to display antibody fragments or proteins at the N-terminus of coat protein pIII?
A: The vector is suitable only for short peptides because displayed proteins longer than 20–30 amino acids can adversely affect the infectivity function of pIII.
Can M13KE be propagated as a plasmid?
A: No, the vector does not carry a plasmid replicon or antibiotic resistance, so it is necessary to propagate the vector as phage, rather than a plasmid
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Customer Reviews
Effective tool for the phage display
It's impressive how the M13KE Phage has offered revolutionary solutions in drug discovery processes, such as developing targeted therapies. Its ability to display peptide libraries on its surface provides a powerful system for identifying potential drug targets.
Worked very well
The M13KE Phage is remarkable in its efficiency and specificity. Its use in phage display techniques has greatly improved our capacity for protein engineering and antibody production.
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