Site-directed mutagenesis

Service Introduction

Site-directed mutagenesis refers to a change of a single base at a specific position in a DNA molecule, usually implying the deliberate alteration of a specific site in the DNA sequence through genetic engineering methods, resulting in specific genotypic and phenotypic changes. This technology has wide applications in life science research, medical treatment, and agricultural production. Through site-directed mutagenesis, genotypes with specific traits or functions can be generated, such as disease models, disease resistance, and salt tolerance. The emergence of site-directed mutagenesis provides a precise and efficient genetic engineering method for the field of life sciences, offering researchers more experimental approaches and research tools.

Service Advantages

▶Experienced: Extensive experience in vector construction, performing over a thousand site-directed mutations annually. Our experienced team can help you quickly complete gene site-directed mutation and construction;

▶Sequence Accuracy: High-performance GS polymerase ensures the accuracy of mutants, avoiding unwanted mutations;

▶Any Site: Using our advanced gene synthesis and mutation technology, mutations can be introduced at any site;

▶Mutations on Long DNA: Optimized technology allows introduction of point mutations, insertion mutations, and deletion mutations on DNA up to 12 kb in length (including the target gene and its vector);

▶One-Stop Solution: Comprehensive upstream and downstream services, including template DNA sequencing, gene synthesis, rapid cloning into expression vectors, custom vector construction, as well as protein expression and purification;

▶Quality Assurance: Ensures that all site-directed mutant vectors are sequence-verified to guarantee product quality.

Service Process

Article Case

An extended set of yeast-based functional assays accurately identifies human disease mutations

academic journal：Method

Assessment of functional effect of missense variants. (A,B) Growth assays on solid media (A) and liquid media (B) for human tyrosyl-tRNA synthetase (YARS) gene variants. G41R and E196K are disease-associated variants; M431L and A339T are non-disease-associated variants. The yeast cells were temperature-sensitive mutants of the yeast TYS1 gene, expressing either wild-type or variant alleles of the YARS gene, or the GFP gene as a control. For solid growth assays, fivefold serial dilutions of yeast cells were spotted onto plates and incubated at 24°C and 36°C for 2 d. For liquid growth assays, approximately 105 cells were inoculated in 100 μL liquid medium and the absorbance at 600 nm was read every 15 min.

Applicable Fields

▶Study of protein functional mechanisms: This is a core tool for analyzing the relationship between protein structure and function. By performing site-directed mutations on key amino acid residues of proteins (such as catalytic active sites, ligand-binding sites, signal peptide regions), changes in protein activity, binding ability, subcellular localization, and other properties after mutation can be observed, clarifying the functional role of specific sites and revealing the molecular mechanisms by which proteins perform their physiological functions.

▶Optimization of industrial proteins and enzyme engineering: Widely applied to enhance the performance of industrial proteins and enzymes. Site-directed mutations on enzymes (such as cellulase, protease, lipase) can improve catalytic efficiency, thermal stability, pH tolerance, or substrate specificity; mutations on functional proteins like antibodies or vaccine antigens can enhance their target-binding ability, immunogenicity, or resistance to degradation, meeting the needs of industrial applications such as biocatalysis and biopharmaceutical production.

▶Microbial metabolic engineering: Supports the efficient construction of microbial cell factories. Site-directed mutations on key genes in microbial metabolic pathways (such as rate-limiting enzyme genes or regulatory factor genes in E. coli or yeast) can control the direction of metabolic flux and increase the synthesis efficiency of target products (like amino acids, organic acids, biofuels, pharmaceutical intermediates); alternatively, mutating microbial transporter genes can enhance product secretion, reduce intracellular toxicity accumulation, and improve fermentation production efficiency.

▶Development of molecular biology tools: Promotes innovation and optimization of new experimental tools. Site-directed mutations on core elements of gene cloning vectors (plasmids, viral vectors) such as promoters, terminators, and restriction sites can improve transcription efficiency, replication stability, or introduce specific restriction sites to simplify the gene cloning process; mutating PCR primer binding regions or gene editing tool recognition sequences can enhance the specificity and practical applicability of these tools.