Mitochondrial Targeting of Catalytic RNAs
Genetic transformation of mitochondria in multicellular eukaryotes has remained inaccessible, hindering fundamental investigations and applications to gene therapy or biotechnology. In this context, we have developed a strategy to target nuclear transgene-encoded RNAs into mitochondria in plants. We describe here mitochondrial targeting of trans-cleaving ribozymes destined to knockdown organelle RNAs for regulation studies and inverse genetics and biotechnological purposes. The design and functional assessment of chimeric RNAs combining the ribozyme and the mitochondrial shuttle are detailed, followed by all procedures to prepare constructs for in vivo expression, generate stable plant transformants, and establish target RNA knockdown in mitochondria.
Key wordsMitochondria Plant Ribozyme RNA knockdown RNA transport Transformation tRNA
This work has been published under the framework of the LABEX [ANR-11-LABX-0057_MITOCROSS] and benefits from a funding from the state managed by the French National Research Agency as part of the “Investments for the future” program. Further support through grants from the French National Research Agency (ANR-06-MRAR-037-02, ANR-09-BLAN-0240-01), the Polish Ministry of Science and Higher Education and the Polish National Science Center is acknowledged. Our projects are also supported by regular funding from the French National Center for Scientific Research (CNRS-UPR2357) and the University of Strasbourg (UdS).
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