Segmental Adeno-Cre Infection: A Technique to Generate Isolated Colorectal Cancer using Genetically Engineered Mouse Models

- Begin by taking an anesthetized conditional mutant mouse with a modified genome carrying specific Cre-recombinase enzyme recognition sequences called LoxP inserts that flank the desired tumor suppressor gene to be mutated. Surgically expose the abdominal cavity. Locate the distal colon and clamp its upper portion. Transanally insert a medical grade tube and advance it into the distal colon. Flush the colon with saline to remove any fecal matter. Replace the tube. Clamp the lower region of the colon for segmentation. Inflate the segment using a suitable cell dissociation solution. This perforates the mucosal barrier and increases the accessibility of intestinal crypt cells. Flush out the dissociation solution.

Next, inject an adenoviral suspension into the colon segment. These engineered viral vectors encode Cre-recombinase enzymes. The virus enters the cell, releases its DNA into the nucleus, and expresses the enzymes. The proteins recognize the LoxP inserts in the host genome and catalyze site specific recombination, thereby inactivating the tumor suppressor gene. This loss of function mutation promotes cancer progression. Suture the incision and allow the mouse to recover. Moni