Enhancer Trap System Essay

Dr. Jim Haseloff A plant biologist working at the Department of Plant Sciences, University of Cambridge. His scientific interests are focused on the engineering of plant morphogenesis, using microscopy, molecular genetics, computational and synthetic biology techniques. Prior to joining the Department of Plant Sciences, Jim served as group leader at MRC Laboratory of Molecular Biology in Cambridge and his group developed advanced imaging techniques and modified fluorescent proteins for efficient use in plants.

Before this, Jim was a research fellow at Harvard Medical School, working on trans-splicing ribozymes. He has also worked at the CSIRO Division of Plant Industry, Canberra, and developed methods for the design of the first synthetic RNA enzymes with novel substrate specificities. Jim is deeply involved with teaching Synthetic Biology at the University of Cambridge, and is very interested in its wider potential as a tool for engineering biological systems and underpinning sustainable technologies.

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He was the one who adapted the GAL4/VP16-UAS system for the construction of enhancer trap lines in Arabidopsis. (Lovell, 2013) GAL4-GFP enhancer trap system It’s a system for GAL4 targeted gene expression. An enhancer trap vector bearing a modified GAL4-VP16 gene was inserted randomly into the Arabidopsis genome by Agro bacterium mediated transformation. Cell specific activation of the GAL4-VP16 gene by cellular enhancer results in activation of a linked GFP gene, allowing simple characterization of expression patterns.

Targeted expression of another gene (X) can be induced by genetic crossing. (Haseloff) Fig1. Shows the two T-DNA constructs. Agrobacterium-mediated transformation Agrobacterium-mediated transformation is used to transform plant cells with a plasmid containing the modified GAL4-VP16 and UAS-GFP genes. The modified GAL4-VP16 gene is positioned adjacent to the Tr border sequence with the truncated (-48) 35S promoter. Transformation results in integration of the T-DNA at random positions into the plant genome.

Proper expression of the GAL4-VP16 gene in these transformed plants requires the fortuitous presence of plant enhancer elements in flanking chromosomal sequences. Expression of the GAL4-VP16 gene then drives the GFP reporter gene. (Haseloff) Selected Arabidopsis GAL4-VP16 enhancer trap lines Confocal micrographs of selected enhancer trap lines, showing root tip specific expression. The roots were counterstained with propidium iodide (red channel). The GAL4-driven GFP signal is shown superimposed (green channel). (Haseloff) Fig. 2 shows roots stained with propidium iodide (red channel) and superimposed (green channel).

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