Chatterjee, Pradeep K.

About two-thirds of the highly conserved genome sequence between human and other vertebrates as divergent as the fish does not code for proteins, is distributed throughout the genome, and mostly located at large distances along the DNA from the start sites of genes. Part of this conserved non-coding DNA plays a role in regulating gene expression from large distances, and is believed to be essential to all vertebrate development. Conservation of gene regulatory function has also been demonstrated in the absence of sequence similarity, suggesting that structural features of DNA can be preserved at those sites despite their different sequence. In fact shape and solvent accessibility of non-coding DNA having similar function appear better preserved evolutionarily than sequence alone. All of these criteria make identification of functional non-coding DNA a real challenge.

We have developed a novel approach to “scan” Bacterial Artificial Chromosomes (BACs) with enhancer-traps to identify functional non-coding DNA in an unbiased and non-targeted manner. BACs are functionalized with enhancer-traps and expressed as transgenes in zebrafish. Using this approach we identified an intron enhancer that is poorly conserved, but required for expression of the Amyloid Precursor Protein (APPb) gene from zebrafish. Identifying functional non-coding DNA this way relies upon expressing genes in their chromosomal contexts in the appropriate tissues and levels reminiscent of their endogenous counterparts.