CROP SCIENCE, plant transformtaion laboratory
Transformation Improvements
Matrix Attachment Regions (MARs)
During the past decade Allen, Spiker, Thompson and Weissinger have developed and patented an NCSU technology which uses DNA elements called Matrix Attachment Regions (MARs) to both improve transformation and to stabilize the expression of transgenic traits. Further improvements are continuing and current research includes the development of a system for predictably and precisely targeting a transgene to a predefined place in the genome. It is hoped that further refinements in the targeting system will allow endogenous genes to be modified and used to replace the native gene using a form of "plant gene therapy".
For users interested in potential commercialization of their transgenic plant, the PTL will help to prevent potential problems that can occur in the interpretation of Intellectual Property (IP) and Freedom to Operate (FTO).
Gene Targeting
A major goal in plant transformation is to control the position of integration of the transgene. We have been able to target genes with high efficiency in NT1 tobacco suspension cells and plants using a novel variation of the Flp/FRT site-specific recombinase system. While we could achieve an extremely high level of site-specific targeting, the resulting targeting event proved to be unstable and and was eventually lost. We have now made several modifications to our system that are designed to improve the stability.
Our current experiments are now focusing on the use of the targeting system in arabidopsis. If we are successful, targeting experiments will be attempted in economically important plants, which typically have large genomes with many repetitive regions. We will then be able to offer the targeting system to the users of the PTL.
DNA Replication
The PTL Director also has basic research programs that are designed to understand the dynamics that occur in the plant nucleus. DNA replication in higher eucaryots remains poorly understood. In 2004 North Carolina State University, Clemson University, and Cold Spring Harbor Laboratories received funding from the NSF Plant Genome Program to research DNA Replication in Arabidopsis and rice. Plants provide a unique opportunity because experiments can be performed with both developing tissues and cultured cells.
The goal of Allen’s group is to map the Matrix Attachment Regions in Arabidopsis and rice cells and to determine whether they function as origins of Plant DNA replication. PhD student Miguel Flores in the Botany Department is currently isolating MAR fragments in order to map their position in the rice and Arabidopsis genomes using microarray analysis. Marina Franco, a Crop Science PhD student is using ChIP to identify DNA fragments bound to the replication complex. These resulting data will be compared to data being produced by our collaborators to gain a more complete knowledge about origins of replication in plants.
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1203 Partners II