De novo synthesis of a chloroplast genome in Nicotiana tabacum

Despite significant progress in chloroplast biotechnology the de novo synthesis of a chloroplast genome hasremained elusive.

The development of strategies to design, assemble and transform artificial chloroplast genomeswould signify a step-change in the field of plant synthetic biology.

Only recently, genome scale DNA synthesisand sequencing has become routine, providing an ideal foundation to revisit the challenge of chloroplast genomesynthesis.

I will join Ralph Bock, a world leader in chloroplast biotechnology, and combine the expertise of hislaboratory with my background in synthetic biology to generate the first Nicotiana tabacum plant with an artificialchloroplast genome.

I will base my strategy on creating and testing multiple different genome designs with varieddegrees of structural and genetic code alterations in respect to the parental genome to identify viable designs.

Iwill develop methods to assess those designs in parallel, permitting the rapid identification and fixing ofproblematic regions in the synthetic genome.

I will proceed to assemble the verified chloroplast genomes in ahierarchical, parallelized manner from large pieces of synthetic DNA.

Finally, I will characterize the phenotype ofthe resulting plants, focusing on understanding the changes in transcription, translation and metabolism.

Themethods developed in this work will provide insights into the design principles of organellar genomes, and providea platform to reinvent and remodel the genetic code of chloroplasts.

This work will thereby establish strategies tore-engineering complex cellular processes such as photosynthesis, or to design entirely new functions such as thegenetic isolation of synthetic DNA in plants.