Overview

Site-directed mutagenesis (SDM) is a powerful tool to study the relationship between protein structure and function. Cloning enzyme sentinel mutagenesis is a commonly used SDM method that involves the introduction of point mutations into a target gene by amplifying the gene with a mutagenic primer. This technique allows precise modification of the sequence of a gene without altering the surrounding DNA, making it ideal for functional studies or for engineering proteins with specific properties. Our cloning enzyme targeted mutagenesis service provides high quality, cost effective and timely results to meet the needs of our clients.

Specific Services We Offer

We offer a range of cloning enzyme targeted mutagenesis services, including

• Single site mutagenesis - Single site mutagenesis involves the introduction of a single mutation in a target gene.

Figure 1. Single site mutagenesis.

• Multiple-site mutagenesis - Multiple-site mutagenesis involves the introduction of two or more mutations in a target gene.

Figure 2. Multiple-site mutagenesis. (Hejlesen E M. 2020)

• Deletion mutagenesis - Deletion mutagenesis is the deletion of one or more nucleotides from a target gene.
• Insertional mutagenesis - Insertional mutagenesis is the insertion of one or more nucleotides into a target gene.
• Codon optimization - This project involves modifying the codon of the target gene to optimize its expression in a specific host system.

Our Methodology

• We use advanced targeted mutagenesis technology to perform our cloning enzyme targeted mutagenesis services. This technology has been extensively validated and optimized to provide high efficiency and precision.
• Our method uses a pair of complementary mutagenesis primers to introduce the desired mutation into the target gene. These primers are designed using proprietary software to ensure optimal primer design for each template sequence.
• After PCR amplification with the mutagenic primers, DpnI digestion is performed to remove the template DNA, leaving only the newly synthesized mutant plasmid. The mutant plasmids are then transformed into E. coli cells for propagation, selection and subsequent analysis.

Sample Requirements

• Template DNA-We require a minimum of 100 ng of high-quality template DNA containing the target gene sequence.
• Mutagenic primers-We require customers to provide their own mutagenic primers, designed using our proprietary primer design software.
• Plasmid vectors-Customers can choose to provide their own plasmid vectors or select from our library of pre-made vectors.

Deliverables

• Mutant plasmids-We provide plasmids containing the desired mutation that can be used for further functional or structural studies.
• Sequencing data-We provide sequencing data to confirm the presence of the desired mutation in the mutagenic plasmid.
• Quality control data-We provide data on the efficiency and accuracy of mutagenesis reactions and other quality control parameters.

Applications

• Functional Studies - Site-directed mutagenesis can be used to study the role of specific residues or structural domains in protein function.
• Structural Biology - Site-directed mutagenesis can be used to generate mutated proteins for crystallography or NMR studies.
• Gene therapy - Site-directed mutagenesis can be used to modify genes for therapeutic purposes, such as correcting an inherited disease or optimizing gene expression.

Why Choose Us?

We offer competitive pricing for our services without compromising on quality or efficiency. Our team of experienced scientists is available to provide expert support and advice throughout the entire process, from primer design to data analysis. If you would like to know more about this service, please feel free to contact us.