Transient Testing of Enzymes Designed for Genome Editing in Maize

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Briggs, Sarah
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Symposium on Undergraduate Research and Creative Expression
Iowa State University Conferences and Symposia

The Symposium provides undergraduates from all academic disciplines with an opportunity to share their research with the university community and other guests through conference-style oral presentations. The Symposium represents part of a larger effort of Iowa State University to enhance, support, and celebrate undergraduate research activity.

Though coordinated by the University Honors Program, all undergraduate students are eligible and encouraged to participate in the Symposium. Undergraduates conducting research but not yet ready to present their work are encouraged to attend the Symposium to learn about the presentation process and students not currently involved in research are encouraged to attend the Symposium to learn about the broad range of undergraduate research activities that are taking place at ISU.

The first Symposium was held in April 2007. The 39 students who presented research and their mentors collectively represented all of ISU's Colleges: Agriculture and Life Sciences, Business, Design, Engineering, Human Sciences, Liberal Arts and Sciences, Veterinary Medicine, and the Graduate College. The event has grown to regularly include more than 100 students presenting on topics that span the broad range of disciplines studied at ISU.

Genetics, Development and Cell Biology

Successful genome editing is associated with the ability to generate double strand breaks (DSB) efficiently at specific chromosomal locations. Recently, enzymes called TALENs (transcription activator like effector nucleases) have been tested for this purpose. TALENs consist of a DNA binding domain and a DNA cleaving or nuclease domain. They function as dimers; two TALEN proteins interact together to produce a DSB in the DNA. The efficiency of TALENs is influenced by the ability to access and bind the target site and of the two TALENs to dimerize. We have assembled a transient assay system in maize to test which TALENs are most effective at generating DSBs. We generated a modified fluorescent protein reporter gene that contains the DNA binding site for the TALENs. When this gene is expressed following DNA bombardment into maize embryos, it is non-functional and no fluorescence is observed. Genes encoding TALENs are then co-bombarded with the reporter gene plasmid. When DSBs occur, DNA repair machinery of the maize cells will repair the reporter gene generating a functional fluorescent protein. The number of fluorescent cells recorded is a measure of TALEN activity. Results from several TALENs and other DSB enzymes, such as RNA-guided cas9 will be discussed.

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