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Validation & phenotyping

Validation & phenotyping

Validation & phenotyping

What is mouse model validation?

Mouse model validation is a check to confirm that the knockout, knock-in, humanized or transgenic mouse model has the correctly engineered allele at the DNA level. Further validation can occur at the level of mRNA expression, protein expression or phenotype.

What is mouse model phenotyping?

Mouse model phenotyping is the assessment of measurable physiological effects caused by genetic changes under specific conditions. This may include immune viability studies, basic morphological and/or behavioural analysis, immune checkpoint target validation, early determination of utility, characterisation under challenge and other pre-clinical mouse research.

Mouse validation and phenotyping

DNA Analysis

At the DNA level, validation of animal models is done by performing DNA analysis to ensure that the desired transgenic modification has been achieved and no unintended mutations have been introduced. Ozgene uses quantitative PCR (qPCR) at specific, unique sites of the targeting vector to ensure that the intended gene has been fully and correctly targeted. Genomic DNA screening is also used to detect any unintended insertional mutations.

mRNA analysis

At the mRNA level, expression of the modified gene is determined by quantifying mRNA as compared to the mRNA levels of the wild-type gene or that of other control genes. This is a valuable check to determine how the genetic modification has altered gene expression. Ozgene uses reverse transcription coupled with qPCR (RT-qPCR) to measure mRNA levels of cells or tissues from mouse models as an indication of relative gene expression.

Protein analysis

Protein expression is also a valuable piece of characterisation data. It relies on the availability of appropriate antibody reagents for analysis by western blotting, flow cytometry or ELISA. These assays provide data regarding changes to protein expression, protein size and the specificity of protein expression across various tissues and cell types. Since protein analysis is downstream of mRNA analysis, it can replace mRNA analysis for validation purposes.

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