Fig. 1
From: A cancer rainbow mouse for visualizing the functional genomics of oncogenic clonal expansion
Engineering cancer rainbow mice. a The current two-step model of intestinal field cancerization. Somatic mutations increase stem cell fitness and lead to the fixation of premalignant clones within crypts (orange). Crypt fission results in duplication of premalignant crypts and the lateral propagation of somatic fields. b The Crainbow expression vector incorporates 4-tandem cassettes downstream of the ubiquitous chicken-β-actin promoter (CAG). The construct confers ubiquitous expression of a membrane-targeted and chemically inducible near-infrared fluorogen-activating peptide (FAP-Mars1) as a control. Cell type specific activation of Cre recombinase mediates recombination at one of three positions through inclusion of three pairs of orthogonal lox sites (LoxN: white triangle, Lox2272: hatched triangle, and LoxP: filled triangle). Single-copy transgene insertion provides a single outcome per cell and barcoding of each genetic fate by fluorescent imaging. c Candidate tumor driver genes are synthesized/PCR amplified and then In-Fusion® cloned into a bicistronic expression cassette. Multi-site Gateway™ cloning is used to directionally clone each pENTR plasmid into a Gateway™-compatible ROSA-targeting vector in a single step21.
