Chloroplast-Based Expression of Recombinant Proteins by Gateway® Cloning Technology
Plastid transformation for the expression of recombinant proteins and entire enzymatic pathways has become a promising tool for plant biotechnology in the past decade. Several improvements of the technology have turned plant plastids into robust and dependable expression platforms for multiple high value compounds. In this chapter, we describe our current methodology based on Gateway® recombinant cloning, which we have adapted for plastid transformation. We describe the steps required for cloning, biolistic transformation, identification, and regeneration of transplastomic plant lines and Western blot analysis.
KeywordsPlastid transformation Gateway® recombinant cloning technology Biolistic transformation Southern blot analysis Western blot analysis
This work was supported by the GLOBVAC program (Project 192510), the Research Council of Norway (RCN), Bioforsk.
- 2.Eibl C, Zou Z, Beck A, Kim M, Mullet J, Koop H-U (1999) In vivo analysis of plastid psbA, rbcL and rpl32 UTR elements by chloroplast transformation: tobacco plastid gene expression is controlled by modulation of transcript levels and translation efficiency. Plant J 19:333–345CrossRefPubMedGoogle Scholar
- 5.Karimi M, Inzé D, Depicker A (2002) GATEWAY™ vectors for Agrobacterium-mediated planttransformation. Trends Plant Sci 7:193–195Google Scholar