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.

Shakes, L.A., Malcolm, T.L., Allen, K.L., De, S., Harewood, K.R., & Chatterjee, P.K. (2008). Context dependent function of APPb Enhancer identified using Enhancer Trap-containing BACs as Transgenes in Zebrafish. Nucleic Acids Research 36: 6237-6248.

Shakes, L. A., Garland, D.M., Srivastava, D.K., Harewood, K.R. and Chatterjee, P. K. (2005). Minimal Cross-recombination between wild type and loxP511 sites in vivo facilitates Truncating Both Ends of Large DNA Inserts in pBACe3.6 and Related Vectors. Nucleic Acids Research. 33: e118.

Chi, X., Chatterjee, P.K., Wilson III, W., Zhang, S-X., DeMayo, F., and Schwartz, R.J. (2005). Complex Cardiac Nkx2-5 Gene Expression Activated By Noggin-Sensitive Enhancers Followed By Chamber-Specific Modules. Proc. Natl. Acad. Sci. (USA) 102: 13490-13495.

Chatterjee P.K., Mukherjee S., Shakes L.A., Wilson III, W., Harewood K.R. & Byrd G. (2004). Selecting Transpositions of a Markerless Transposon Using Phage P1 Headful Packaging: New Transposons for Functionally Mapping Long Range Regulatory Sequences in BACs. Analytical Biochemistry 335: 305-315.

Chatterjee P.K., Shakes L.A., Srivastava D.K., Garland D.M., Harewood K.R., Moore K.J. & Coren J.S. (2004). Mutually Exclusive Recombination of Wild Type and Mutant loxP Sites in vivo Facilitates Transposon-Mediated Deletions from Both Ends of Genomic DNA in PACs. Nucleic Acids Research 32: 5668-5676.

Gilmore, R.C., Baker Jr., J., Dempsey, S., Marchan, R., Corprew Jr., R.N.L., Byrd, G., Maeda, N., Smithies, O., Bukoski, R.D., Harewood, K.R. & Chatterjee, P.K. (2001). Using PAC nested-deletions to order contigs and microsatellite markers at the high repetitive sequence containing Npr3 gene locus. Gene 275:65-72.

Chatterjee, P.K., and Briley, L.P. (2000). Analysis of a Clonal Selection Event During Transposon-Mediated Nested-Deletion Formation in Rare BAC and PAC Clones. Analytical Biochemistry 285: 121-126.

Chatterjee, P.K., Yarnall, D.P., Haneline, S.A., Godlevski, M.M., Thornber , S.J., Robinson, P.S., Davies, H.E., White, N.J., Riley, J.H. and Shepherd, N.S. (1999). Direct Sequencing of Bacterial and P1 Artificial Chromosome Nested-deletions for Identifying Position-Specific Single Nucleotide Polymorphisms. Proc. Natl. Acad. Sci. (USA) 96: 13276-13281.

Chatterjee, P.K. and Coren, J.S. (1997). Isolating Large Nested Deletions in Bacterial and P1 Artificial Chromosomes by in vivo P1 Packaging of Products of Cre-catalysed Recombination Between the Endogenous and a Transposed loxP Site. Nucleic Acids Research 25: 2205-2212.

Chatterjee, P.K. and Sternberg, N. L. (1995). A General Genetic Approach in E.coli for Determining the Mechanism(s) of Action of Tumoricidal Agents: Application to DMP 840, a tumoricidal agent. Proc. Natl. Acad. Sci. (USA) 92, 8950-8954.

Chatterjee, P.K.,   Pruzan, R.  and Flint, S.J.  (1993). Purification of an active TATA-binding protein-containing factor using a Monoclonal Antibody that recognizes the Human TATA-Binding Protein. Protein Expression and Purification. 4, 445-455.

Pruzan, R., Chatterjee, P.K. and Flint, S.J. (1992). Specific Transcription from the Adenovirus E2E Promoter by RNA Polymerase III requires a subpopulation of TFIID. Nucleic Acids Research 20, 5705-5712.

Chatterjee, P.K. and Flint, S.J. (1987). The Adenovirus type 2 Endopeptidase:  An Unusual Phosphoprotein Enzyme Matured by Autocatalysis. Proc. Natl. Acad. Sci. (USA) 84, 714-718.