Research Agenda Examine mouse anatomy

Choose organ/tissue for structural/histological examination

Extract DNA from mouse tissue sample

Amplify DNA in PCR to identify a gene of interest (genotype)

Research Agenda Run DNA Gel Electrophoresis to visualize genotyping results

Use bacterial transformation to express and replicate a gene of interest for in vitro studies

Check your transformation to verify gene presence using an enzyme digest assay

Examine protein localization in tissue using ImmunoHistoChemistry (IHC)

Mouse Anatomy Secure mouse on styrofoam blocks with pins Make a ventral midline incision, then cut across abdominal cavity, diaphragm and upper

chest. You will need to cut through the sternum and ribs to expose heart and lungs

Mouse Anatomy Locate the heart, lungs, diaphragm, liver, gal bladder, spleen, stomach, small intestines,

cecum, large intestine, bladder, kidneys Choose one organ to harvest for further histological examination: heart, brain, kidney, liver

Tissue Fixation By fixation we intend to:• (1) conserve the tissue from autolysis and

bacterial attack• (2) prevent the loss of cellular constituents• (3) increase optical differentiation of

cellular structures• (4) increase tissue consistency, in order to

facilitate their going through the other steps of the technique – especially slicing

How it works: Paraformaldehyde will crosslink primary amino

groups in proteins with other nearby nitrogen atoms in protein or DNA through a –CH2- linkage

A solution of 4% formaldehyde fixes pathology tissue specimens at about one mm per hour at room temperature

Depending on duration of exposure and temperature, fixation may be reversible or irreversible

Tail Snip Cut a < 5 mm tail section and

submerge it into DNA extraction buffer

Proteinase K enzyme buffer will digest tissue overnight and we will harvest the DNA tomorrow for further examination