Point Mutation Mouse    

Point mutant mice are gene knockout mouse models in which one or more nucleotides are replaced by mutant nucleotides in the mouse genome. This may result in amino acid changes within the protein sequence, and transcription terminated, thereby significantly affecting the normal function of the protein. Point mutation mouse models are widely used to study the role of specific nucleotides or amino acids in genetic elements or proteins, as well as to mimic human genetic diseases. CRISPR / Cas9 technology provides a very efficient tool to generate point mutations in mouse models, to generate precise nucleotides substitutions at target sites.

The CRISPR/Cas9 Platform^CB, one of the leading gene-editing biology companies, has successfully obtained >200 mouse models with very high efficiency and success rate by using CRISPR/Cas9-mediated genome editing technology. We are committed to providing you with high-quality CRISPR/Cas9 gene-editing services, including custom CRISPR/Cas9 design strategy, and trusted verification methods. We guarantee delivery of at least 2 founders or 3 F1 mice and complete within 3-4 months.

Workflow of CRISPR-Cas9-Mediated Pointed Mutation Mouse Services

• Point Mutation Strategy Design

Based on the genes you are interested in, we will provide you with a comprehensive and efficient point mutation strategy to maximize Cas9 efficiency, including the selection of appropriate target sites, gRNA design and synthesis, single-stranded donor oligonucleotide (ssDNA) design and synthesis, vector construction, PCR primers for genotypic identification.

• CRISPR Vector Construction

Synthesize gRNA, Cas9 mRNA and ssDNA (carried the desired mutation), construct vector.

• Injection into Mouse Embryos

Inject gRNA designed for a specific site of the target gene, Cas9 mRNA and ssDNA (carried the desired mutation) simultaneously into the mouse embryos. Typically, we will inject as many embryos as needed to fulfill the guarantee.

• Genotyping of Founder Mice

Embryos are transplanted into pseudopregnant mice to produce founder mice. The founders are verified and screened by PCR and sequencing analysis to reveal any mutation at targeted sites.

• Breeding of Founders to Obtain F1 Mice

According to your needs, we will decide whether or not to breed the founder mice. Normally, the founders will be mated with wildtype mice of matched genetic background to generate their offspring. By genotyping of their offspring, we will select the F1 mice bearing the point mutation allele.

Technical Details

By simultaneously injecting gRNA, Cas9 mRNA and ssDNA into mouse fertilized eggs, we introduced defined genomic modifications in the fertilized egg genome at a lower cost and in a shorter time. Cas9 binds to gRNA, and causes targeted double-strand breaks (DSBs) in genomic DNA, ssDNA acts as a DNA repair template that will lead to one or more nucleotides mutated at the target site during homologous directed repair (HDR).

Typically, we develop CRISPR-mediated knockout mice in C57BL/6 and FVB, but we can use other mouse strains according to your requirements.

Benefits

• One-stop service from strategy design to F1 mice
• Faster turnaround and lower cost
• F1 breeding to ensure germline transmission
• Timely report on project progress

Application

• Study the functions of specific mutant genes/proteins
• Analysis of the role of non-coding regions and regulatory factors
• Human disease model
• Screening and verification of disease-causing mutations
• Study resistant mutations
• Research Drugs - Antibody Affinity
• Antibody screening/validation
• Preclinical safety and toxicity studies

CRISPR/Cas9 Platform^CB has national labs, a team of scientists from a world-leading university, and has extensive experience in editing CRISPR/Cas genes. Tell us about the genes you are interested in and we will provide you with a professional CRISPR/Cas point mutation mouse model strategy. During the implementation of the project, we will promptly inform you of the progress, and at the end provide a final report detailing the test strategy and verification method. we will meet your specific requirements.