Analysis of Expression and Imprinting of p57Kip2 from a Modified BAC Transgene

Rosalind John
Wellcome/CRC Institute

p57Kip2 is a maternally-expressed, imprinted gene which encodes a transcript of approximately 1.4 kb and spans a 3 kb genomic region. It lies within the established imprinted domain on distal chromosome 7 where a number of imprinted genes are clustered. This region is syntenic to human chromosome 11p15 where the gene order and imprinting are conserved. In humans, mutations of this region are associated with Beckwith-Wiedemann Syndrome (BWS).

We have isolated and characterized bacterial artificial chromosomes (BACs) which span approximately 320 kb around the p57Kip2 locus. To identify the control cis-elements involved in expression and imprinting of p57Kip2 we are testing the ability of these BACs to imprint in the mouse. Using a recently described technique which allows the modification of E.coli-based genomic clones, we have homologously recombined a two-color reporter gene into the 3'-UTR of p57Kip2 and generated transgenic mouse lines by pronuclear injection. One transgene directs a strikingly restricted expression profile while a second, overlapping transgene appears to contain all the enhancers required for appropriate expression. Preliminary analysis of transgenic lines suggests that key regulatory elements for the imprinted expression of p57Kip2 lie at a distance from the gene. Implications for BWS will be discussed.