journal of Histology & Histopathology

Molecular Biology and Genetic Engineering

ISSN 2053-5767
Review

Long range gene-regulatory sequences identified by transgenic expression of bacterially-engineered enhancer-trap BACs in zebrafish

Hope M. Wolf1,2, Kevin O. Nyabera1, Katya K. De La Torre1, Mugtaba A. Eltayeb1, Oladoyin Iranloye1, Leighcraft A. Shakes1, Charles Hatcher1, Derek C. Norford1 and Pradeep K. Chatterjee1*

*Correspondence: Pradeep K. Chatterjee pchatterjee@nccu.edu

1. Julius L. Chambers Biomedical/Biotechnology Research Institute and Department of Chemistry, North Carolina Central University, 1801 Fayetteville Street, Durham, NC 27707, USA.

Author Affiliations

2. Department of Chemistry, University of North Carolina at Chapel-Hill, Chapel-Hill, NC 27599, USA.

Abstract

Large pieces of DNA from the chromosomes of numerous organisms, including the human, are faithfully propagated in bacteria as large extra-chromosomal plasmids known as Bacterial Artificial Chromosomes (BACs). Because they represent tiny contiguous pieces of the chromosome, BACs are ideally suited for expression of genes in their chromosomal contexts. Genes in BACs need to be functionalized with reporter genes and other sequences that allow easy monitoring, stable maintenance and propagation of the DNA in the new host organism. BAC DNA can be altered within its bacterial host in several ways. One approach uses Tn10 mini-transposons to introduce exogenous DNA into BACs for a variety of purposes. The random insertions of Tn10 transposons carrying lox sites have helped position mammalian cellselectable antibiotic resistance genes, enhancer-traps and inverted repeat end-sequences of the vertebrate transposon Tol2 precisely at the ends of genomic DNA inserts in BACs. Functional identification of generegulatory elements through reporter gene expression and BAC DNA integration into zebrafish or mouse chromosomes have been facilitated with such retrofitting. The methodology has been used extensively to dissect the regulation of the Amyloid Precursor Protein (appb) gene in zebrafish. Functional identification of long-range regulatory sequences of appb has provided important clues for regulation of the APP gene in humans.

Keywords: Enhancer-trap BACs, BAC transgenesis, zebrafish transgenesis, regulation by E4BP4/NFIL3, reason for AD connection to circadian clock, alzheimer's disease

ISSN 2053-5767
Volume 2
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