APPLICATIONS OF TECHNOLOGY:
- Industrial biotechnology
- DNA mutation repair
- Genetics in organelles
- Efficient, homoplasmic integrations in the mitochondrial genome
- Genetic change of the mtDNA without a selection marker
Targeted genome engineering for mitochondria and other organelles has important implications in fungal, plant, algal and human cells and tissues. Unlike the nuclear genome, mitochondrial DNA (mtDNA) exists in high copy numbers in a single cell, thereby serving as an attractive target for high-level, heterologous gene expression in biotechnology hosts, such as yeast. However, this same feature presents a challenge for introducing homogenous genetic changes, which this invention provides a method for overcoming.
Researchers at Lawrence Berkeley National Laboratory’s Joint BioEnergy Institute (JBEI) have developed a technology for making homogenous sequence modifications (edits or integration) into the mitochondrial genome of eukaryotic cells. Previous gene editing tools have only focused on the nuclear genome, while the JBEI invention allows for efficient, reliable genetic engineering in the mitochondrial genome. Mitochondrial genome exists in many (hundreds) of copies per cell, making an ideal target for many applications (e.g. high expression of metabolic pathways), but homoplasmic edits are difficult to achieve in the absence of a selection against unedited copies.
The JBEI technology ensures the import of accessory RNA and DNA needed for genetic modifications. Cas9, a targeted RNA-guided DNA endonuclease, is imported into the mitochondria. By using an endonuclease that specifically targets un-edited DNA loci, the scientists achieved efficient, homoplasmic integrations in the mitochondrial genome. Integrations included modifications to native mitochondrial genes to improve and modify host metabolism, and the introduction of heterologous metabolic pathways for bioproduction.
Existing technologies for modifying mtDNA require a selection, generally as complementation of a mutated gene, to achieve homoplasmic integration. This technology allows for any genetic change of the mtDNA without a selection marker or other sequence constraints. The invention also provides novel techniques for importing nucleic acids (DNA and RNA) needed for Cas9-based gene editing into the mitochondrial matrix. This invention therefore allows for efficient, reliable genetic engineering in the mitochondria, which has not been previously possible.
Amand Reider Apel
DEVELOPMENT STAGE: Proven principle.
STATUS: Patent pending. Available for licensing or collaborative research.
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