Nanopore sequencing identifies a new Tyr::CreERT2 allele circulating in existing mouse stocks

Abstract

The Tyr::CreERT2 transgenic mouse lines are essential tools for conditional gene manipulation in melanocytes and are widely used in melanoma research. Two independent lines are in common use: the Bosenberg line and the Larue line. Precise knowledge of transgene integration sites is critical for designing complex genetic crosses, yet the integration sites for these lines have only recently been characterised by whole genome sequencing. Here we report that a Tyr::CreERT2 mouse stock routinely used in BrafCA;Ptenflox melanoma models carries a previously undescribed integration on Chromosome 1, distinct from the previously reported Chromosome 2 integration. Using long-read nanopore sequencing, we mapped the integration to an intergenic locus between Alppl2 and Alpi, revealing a 2,430 bp genomic deletion at the insertion site. We developed position-specific junction PCR and qPCR assays to genotype this allele and confirmed that offspring are born at Mendelian ratios. Given that this stock is associated with elevated spontaneous melanoma penetrance, accurate characterisation of this allele has direct implications for the many laboratories employing this widely distributed melanoma model. SIGNIFICANCEThis study identifies a previously undescribed Tyr::CreERT2 transgene integration on Chromosome 1 in JAX strain 013590, a line widely used across the melanoma research community. Accurate characterisation of this allele has direct practical implications for the many laboratories that rely on this model for studies of melanoma initiation and progression.