Advertisement

Tools and Techniques for Chloroplast Transformation of Chlamydomonas

  • Saul Purton
Part of the Advances in Experimental Medicine and Biology book series (volume 616)

Abstract

The chloroplast organelle of plant and algal cells contains its own genetic system with a genome of a hundred or so genes. Stable transformation of the chloroplast was first achieved in 1988, using the newly developed biolistic method of DNA delivery to introduce cloned DNA into the genome of the green unicellular alga Chlamydomonas reinhardtii. Since that time there have been significant developments in chloroplast genetic engineering using this versatile organism, and it is probable that the next few years will see increasing interest in commercial applications whereby high-value therapeutic proteins and other recombinant products are synthesized in the Chlamydomonas chloroplast. In this chapter I review the basic methodology of chloroplast transformation, the current techniques and applications, and the future possibilities for using the Chlamydomonas chloroplast as a green organelle factory.

Keywords

Selectable Marker Foreign Gene Chloroplast Gene Chloroplast Transformation Chloroplast Gene Expression 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Sugiura M. The chloroplast genome. Essays Biochem 1995; 30:49–57.PubMedGoogle Scholar
  2. 2.
    Bogorad L. Engineering chloroplasts: An alternative site for foreign genes, reactions and products. Trends Biotechnol 2000; 18:257–263.PubMedCrossRefGoogle Scholar
  3. 3.
    Maliga P. Plastid transformation in higher plants. Annu Rev Plant Biol 2004; 55:289–313.PubMedCrossRefGoogle Scholar
  4. 4.
    Franklin SE, Mayfield SP. Prospects for molecular farming in the green alga Chlamydomonas. Curr Opin Plant Biol 2004; 7:159–165.PubMedCrossRefGoogle Scholar
  5. 5.
    Boynton JE, Gillham NW, Harris EH et al. Chloroplast transformation in Chlamydomonas with high velocity microprojectiles. Science 1988; 240:1534–1538.PubMedCrossRefGoogle Scholar
  6. 6.
    Goidschmidt-Clermont M. Transgenic expression of aminoglycoside adenine transferase in the chloroplast: A selectable marker for site-directed transformation of Chlamydomonas. Nucleic Acids Res 1991; 19:4083–4089.CrossRefGoogle Scholar
  7. 7.
    Lister DL, Bateman JM, Purton S et al. DNA transfer from chloroplast to nucleus is much rarer in Chlamydomonas than in tobacco. Gene 2003; 316:33–38.PubMedCrossRefGoogle Scholar
  8. 8.
    Mayfield SP, Franklin SE, Lerner RA. Expression and assembly of a fully active antibody in algae. Proc Natl Acad Sci USA 2003; 100:438–442.PubMedCrossRefGoogle Scholar
  9. 9.
    Boynton JE, Gillham NW. Chloroplast transformation in Chlamydomonas. Methods Enzymol 1993; 217:510–536.PubMedCrossRefGoogle Scholar
  10. 10.
    Erickson JM. Chloroplast transformation: Current results and future prospects. In: Ort DR, Yocum CF, eds. Oxygenic Photosynthesis: The Light Reactions. Dordrecht: Kluwer Academic Publishers, 1996:589–619.Google Scholar
  11. 11.
    Goidschmidt-Clermont M. Chloroplast transformation and reverse genetics. In: Rochaix JD, Goidschmidt-Clermont M, Merchant S, eds. The Molecular Biology of Chloroplasts and Mitochondria in Chlamydomonas. Dordrecht: Kluwer Academic Publishers, 1998:139–149.Google Scholar
  12. 12.
    Taylor NJ, Fauquet CM. Microparticle bombardment as a tool in plant science and agricultural biotechnology. DNA Cell Biol 2002; 21:963–977.PubMedCrossRefGoogle Scholar
  13. 13.
    Sanford JC. The biolistic process. Trends Biotechnol 1998; 6:299–302.CrossRefGoogle Scholar
  14. 14.
    Zumbrunn G, Schneider M, Rochaix JR. A simple particle gun for DNA-mediated cell transformation. Technique 1989; 1:204–216.Google Scholar
  15. 15.
    Kindle KL, Richards KL, Stern DB. Engineering the chloroplast genome: Techniques and capabilities for chloroplast transformation in Chlamydomonas reinhardtii. Proc Natl Acad Sci USA 1991; 88:1721–1725.PubMedCrossRefGoogle Scholar
  16. 16.
    Rochaix JD. Chlamydomonas reinhardtii as the photosynthetic yeast. Annu Rev Genet 1995; 29:209–230.PubMedCrossRefGoogle Scholar
  17. 17.
    Walker TL, Collet C, Purton S. Algal transgenics in the genomic era. J Phycol 2005; 41:1077–1093.CrossRefGoogle Scholar
  18. 18.
    Cerutti H, Johnson AM, Boynton JE et al. Inhibition of chloroplast DNA recombination and repair by dominant negative mutants of Escherichia coli RecA. Mol Cell Biol 1995; 16:3003–3011.Google Scholar
  19. 19.
    Dauvillee D, Hilbig L, Preiss S et al. Minimal extent of sequence homology required for homologous recombination at the psbA locus in Chlamydomonas reinhardtii chloroplasts using PCR-generated DNA fragments. Photosynth Res 2004; 79:219–224.PubMedCrossRefGoogle Scholar
  20. 20.
    Fischer N, Stampacchia O, Redding K et al. Selectable marker recycling in the chloroplast. Mol Gen Genet 1996; 251:373–380.PubMedCrossRefGoogle Scholar
  21. 21.
    Künstner P, Guardiola A, Takahashi Y et al. A mutant strain of Chlamydomonas reinhardtii lacking the chloroplast photosystem II psbI gene grows photoautotrophically. J Biol Chem 1995; 270:9651–9654.PubMedCrossRefGoogle Scholar
  22. 22.
    Guergova-Kuras M, Boudreaux B, Joliot A et al. Evidence for two active branches for electron transfer in photosystem I. Proc Natl Acad Sci USA 2001; 98:4437–4442.PubMedCrossRefGoogle Scholar
  23. 23.
    Bateman JM, Purton S. Tools for chloroplast transformation in Chlamydomonas: Expression vectors and a new dominant selectable marker. Mol Gen Genet 2000; 263:404–410.PubMedCrossRefGoogle Scholar
  24. 24.
    Rochaix JD. Functional analysis of plastid genes through chloroplast reverse genetics in Chlamydomonas. In: Larkum AWD, Douglas SE, Raven JA, eds. Photosynthesis in Algae. Dordrecht: Kluwer Academic Publishers, 2003:83–94.Google Scholar
  25. 25.
    Xiong L, Sayre RT. Engineering the chloroplast encoded proteins of Chlamydomonas. Photosynth Res 2004; 80:411–419.PubMedCrossRefGoogle Scholar
  26. 26.
    Hauser CR, Gillham NW, Boynton JE. Regulation of chloroplast translation. In: Rochaix JD, Goldschmidt-Clermont M, Merchant S, eds. The Molecular Biology of Chloroplasts and Mitochondria in Chlamydomonas. Dordrecht: Kluwer Academic Publishers, 1998:197–217.Google Scholar
  27. 27.
    Sakamoto W, Kindle KL, Stern DB. In vivo analysis of Chlamydomonas chloroplast petD gene expression using stable transformation of beta-glucuronidase translational fusions. Proc Natl Acad Sci USA 1993; 90:497–501.PubMedCrossRefGoogle Scholar
  28. 28.
    Komine Y, Kikis E, Schuster G et al. Evidence for in vivo modulation of chloroplast RNA stability by 3′-UTR homopolymeric tails in Chlamydomonas reinhardtii. Proc Natl Acad Sci USA 2002; 99:4085–4090.PubMedCrossRefGoogle Scholar
  29. 29.
    Franklin S, Ngo B, Efuet E et al. Development of a GFP reporter gene for Chlamydomonas reinhardtii chloroplast. Plant J 2002; 30:733–744.PubMedCrossRefGoogle Scholar
  30. 30.
    Mayfield SP, Schultz J. Development of a luciferase reporter gene, luxCt, for Chlamydomonas reinhardtii chloroplast. Plant J 2004; 37:449–458.PubMedCrossRefGoogle Scholar
  31. 31.
    Choquet Y, Zito F, Wostrikoff K et al. Cytochrome f translation in Chlamydomonas chloroplast is autoregulated by its carboxyl-terminal domain. Plant Cell 2003; 15:1443–1454.PubMedCrossRefGoogle Scholar
  32. 32.
    Franklin JL, Zhang J, Redding K. Use of aminoglycoside adenyltransferase translational fusions to determine topology of thylakoid membrane proteins. FEBS Lett 2003; 536:97–100.PubMedCrossRefGoogle Scholar
  33. 33.
    Sun M, Qian K, Su N et al. Foot-and-mouth disease virus VP1 protein fused with cholera toxin B subunit expressed in Chlamydomonas reinhardtii chloroplast. Biotechnol Lett 2003; 25:1087–1092.PubMedCrossRefGoogle Scholar
  34. 34.
    Fukusaki EI, Nishikawa T, Kato K et al. Introduction of the archaebacterial geranylgeranyl pyrophosphate synthase gene into Chlamydomonas reinhardtii chloroplast. J Biosci Bioeng 2003; 95:283–287.PubMedGoogle Scholar
  35. 35.
    Cheng Q, Day A, Dowson-Day M et al. The Klebsiella pneumoniae nitrogenase Fe protein gene (nifH) functionally substitutes for the chlL gene in Chlamydomonas reinhardtii. Biochem Biophys Res Comm 2005; 329:966–975.PubMedCrossRefGoogle Scholar
  36. 36.
    Su ZL, Qian KX, Tan CP et al. Recombination and heterologous expression of allophycocyanin in the chloroplast of Chlamydomonas reinhardtii. Acta Biochimica et Biophysica Sinica 2005; 37:709–712.PubMedCrossRefGoogle Scholar
  37. 37.
    Eberhard S, Drapier D, Wollman FA. Searching limiting steps in the expression of chloroplast-encoded proteins: Relations between gene copy number, transcription, transcript abundance and translation rate in the chloroplast of Chlamydomonas reinhardtii. Plant J 2002; 31:149–160.PubMedCrossRefGoogle Scholar
  38. 38.
    Barnes D, Franklin S, Schultz et al. Contribution of 5′ and 3′ untranslated regions of plastid mRNAs to the expression of Chlamydomonas reinhardtii chloroplast genes. Mol Gen Genomics 2005; 274:625–636.CrossRefGoogle Scholar
  39. 39.
    Lössl A, Bohmert K, Harloff H et al. Inducible trans-activation of plastid transgenes: Expression of the R. eutropha phb operon in transplastomic tobacco. Plant Cell Physiol 2005; 46:1462–1471.PubMedCrossRefGoogle Scholar
  40. 40.
    Mühlbauer SK, Koop HU. External control of transgene expression in tobacco plastids using the bacterial lac repressor. Plant J 2005; 43:941–946.PubMedCrossRefGoogle Scholar
  41. 41.
    Apt KE, Behren PW. Commercial developments in microalgal biotechnology. J Phycol 1999; 35:215–226.CrossRefGoogle Scholar
  42. 42.
    Franklin SE, Mayfield SP. Recent developments in the production of human therapeutic proteins in eukaryotic algae. Expert Opin Biol Ther 2005; 5:225–235.PubMedCrossRefGoogle Scholar
  43. 43.
    Ishukura K, Takaoka Y, Kato K et al. Expression of a foreign gene in Chlamydomonas reinhardtii chloroplast. J Biosci Bioeng 1999; 87:307–314.CrossRefGoogle Scholar
  44. 44.
    Minko I, Holloway SP, Nikaido S et al. Renilla luciferase as a vital reporter for chloroplast gene expression in Chlamydomonas. Mol Gen Genet 1999; 262:421–425.PubMedCrossRefGoogle Scholar

Copyright information

© Landes Bioscience and Springer Science+Business Media 2007

Authors and Affiliations

  • Saul Purton
    • 1
  1. 1.Algal Research Group, Department of BiologyUniversity College LondonLondonUK

Personalised recommendations