How can you determine the genotype of a male mouse? - briefly
Extract DNA from a tissue sample (e.g., tail tip), amplify the target locus by PCR, and identify the alleles through gel electrophoresis, restriction analysis, or DNA sequencing. Alternatively, infer the genotype by examining the segregation of traits in progeny from a controlled cross with a known genotype.
How can you determine the genotype of a male mouse? - in detail
Determining the genetic composition of a male laboratory mouse requires a combination of molecular, phenotypic, and breeding approaches.
Molecular analysis provides the most direct evidence. Extract genomic DNA from a tissue sample such as tail snip or ear punch. Perform polymerase chain reaction (PCR) with primers specific to the locus of interest. Analyze amplified products by agarose‑gel electrophoresis or capillary electrophoresis to detect size differences, insertions, deletions, or presence of transgenes. For single‑nucleotide variants, apply Sanger sequencing or targeted next‑generation sequencing; compare the obtained sequence to the reference genome to identify alleles. Quantitative PCR (qPCR) or digital droplet PCR can measure copy number of engineered constructs.
Phenotypic assessment supplements molecular data when the target gene influences observable traits. Record coat color, ear morphology, eye pigmentation, or behavioral markers known to be linked to the genotype. Use standardized scoring systems to reduce observer bias. When the phenotype is recessive, cross the male with a homozygous recessive female and examine the F1 offspring; the presence or absence of the trait confirms the male’s allele status.
Breeding strategies verify inheritance patterns. Set up a test cross with a mouse of known genotype. Analyze the genotypes of the progeny using PCR or phenotype scoring. A 1:1 segregation of alleles indicates heterozygosity in the sire; a uniform genotype suggests homozygosity. Backcross the male to a reference strain and genotype the resulting litter to confirm allele transmission.
A practical workflow:
- Collect tissue, isolate DNA, quantify concentration.
- Design or obtain locus‑specific primers; include control primers for a housekeeping gene.
- Run PCR; verify product size on a gel.
- If necessary, purify PCR product and submit for sequencing.
- Interpret sequence data against reference; note heterozygous peaks or indels.
- Corroborate molecular findings with phenotypic observation.
- Perform a test cross; genotype offspring to confirm allele status.
By integrating PCR‑based genotyping, sequencing verification, phenotypic scoring, and controlled breeding, researchers obtain a reliable determination of a male mouse’s genetic makeup.