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A galinstan expansion femtosyringe for microinjection of eukaryotic organelles and prokaryotes

Nature Biotechnology volume 17, pages 906–909 (1999)Cite this article

Abstract

A galinstan expansion femtosyringe enables femtoliter to attoliter samples to be introduced into prokaryotes and subcellular compartments of eukaryotes. The method uses heat-induced expansion of galinstan (a liquid metal alloy of gallium, indium, and tin) within a glass syringe to expel samples through a tip diameter of about 0.1 μm. The narrow tip inflicts less damage than conventional capillaries, and the heat-induced expansion of the galinstan allows fine control over the rate of injection. We demonstrate injection of Lucifer Yellow and Lucifer Yellow–dextran conjugates into cyanobacteria, and into nuclei and chloroplasts of higher organisms. Injection of a plasmid containing the bla gene into the cyanobacterium Phormidium laminosum resulted in transformed ampicillin-resistant cultures. Green fluorescent protein was expressed in attached leaves of tobacco and Vicia faba following injection of DNA cantaining its gene into individual chloroplasts.

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Figure 1: Scanning electron microscope images showing (A) a conventional microelectrode tip (diameter ~0.7 μm) and (B) a GEF tip (diameter ~0.1 μm).
Figure 2: Confocal laser scanning microscope images of Lucifer Yellow injected into cells and organelles.
Figure 3: GFP expression in chloroplasts following GEF injection of DNA.
Figure 4: (A) Microinjection of Lucifer Yellow (shown in green) into a single chloroplast of a Vicia faba mesophyll cell.

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References

  1. Kempers, R. & van Bel, A.J.E. Symplasmic connections between sieve element and companion cell in the stem phloem of Vicia faba L. have a molecular exclusion limit of at least 10 kDa. Planta 201, 195–201 (1997).

    Article  CAS  Google Scholar 

  2. Kamei, Y. et al. A CBP integrator complex mediates transcriptional activation and AP-1 inhibition by nuclear receptors. Cell 85, 403– 414 (1996).

    Article  CAS  Google Scholar 

  3. Oka, M.T. Arai, T. & Hamaguchi, Y. Microinjection of the monoclonal anti-tubulin antibody YL1/2 inhibits cleavage of sand dollar eggs. Cell Struct. Func. 15, 373–378 ( 1990).

    Article  CAS  Google Scholar 

  4. Höner, B., Citi, S., Kendrick-Jones, J. & Jockusch, B.M. Modulation of cellular morphology and locomotory activity by antibodies against myosin. J. Cell. Biol. 107, 2181– 2189 (1998).

    Article  Google Scholar 

  5. Rose, D.W., McCabe, G., Fermisco, J.R. & Adler, M. Expression of c-fos and AP-1 activity in senescent human fibroblasts is not sufficient for DNA synthesis. J. Cell. Biol. 119, 1405–1411 (1992).

    Article  CAS  Google Scholar 

  6. Staiger, C.J. et al. Microinjected profilin affects cytoplasmic streaming in plant cells by rapidly depolymerizing actin microfilaments. Curr. Biol. 4, 215–219 (1994).

    Article  CAS  Google Scholar 

  7. Walton, P.A., Gould, S.J., Rachubinski, R.A., Subramani, S. & Feramisco J.R. Transport of microinjected alcohol oxidase from Pichia pastoris into vesicles in mammalian cells: involvement of the peroxisomal targeting signal. J. Cell. Biol. 118, 499–508 (1992).

    Article  CAS  Google Scholar 

  8. Kost, B., Galli, A., Potrykus, I. & Neuhaus, G. High efficiency transient and stable transformation by optimized DNA microinjection into Nicotiana tabacum protoplasts. J. Exp. Bot. 46, 1157–1167 (1995).

    Article  CAS  Google Scholar 

  9. Nguyen, L., Lucas, W.J., Ding, B. & Zaitlin, M. Viral RNA trafficking is inhibited in replicase-mediated resistant transgenic tobacco plants. Proc. Natl. Acad. Sci. USA 93, 12643– 12647 (1996).

    Article  CAS  Google Scholar 

  10. Heinzel, T. et al. A complex containing N-CoR, mSin3 and histone deacetylase mediates transcriptional repression. Nature 387, 43–48 (1997).

    Article  CAS  Google Scholar 

  11. Van der Schoot, C. Determinants of xylem-to-phloem transfer in tomato. Thesis, Univ. Utrecht (1989).

  12. Knoblauch, M. & van Bel, A.J.E. Sieve tubes in action. Plant Cell 10, 35–50 ( 1998).

    Article  CAS  Google Scholar 

  13. Hibberd, J.M., Linley, P.J., Khan, M.S. & Gray, J.C. Transient expression of green fluorescent protein in various plastid types following microprojectile bombardment. Plant J. 16, 627– 632 (1998).

    Article  CAS  Google Scholar 

  14. Golds, T., Maliga, P. & Koop, H.U. Stable plastid transformation in PEG-treated protoplasts of Nicotiana tabacum. Bio/Technology 11, 95–97 (1993).

    CAS  Google Scholar 

  15. Haseloff, J., Siemering, K.R., Prasher, D.C. & Hodge, S. Removal of a cryptic intron and subcellular localization of green fluorescent protein are required to mark transgenic Arabidopsis plants brightly. Proc. Natl. Acad. Sci. USA 94, 2122– 2127 (1997).

    Article  CAS  Google Scholar 

  16. Grebenok, R.J. et al. Green-fluorescent protein fusions for efficient characterisation of nuclear targeting. Plant J. 11, 573– 586 (1997).

    Article  CAS  Google Scholar 

  17. Köhler, R.H., Hansen, M.R. & Wildman, S.G. Plastid interconnections imaged by fluorescence and phase contrast. Trends Cell. Biol. 7, 392 (1997).

    Google Scholar 

  18. Köhler, R.H., Cao, J., Zipfel, W.R., Webb, W.W. & Hanson, M.R. Exchange of protein molecules through connections between higher plant plastids. Science 276, 2039–2042 (1997).

    Article  Google Scholar 

  19. Svab, Z. & Maliga, P. High frequency plastid transformation in tobacco by selection for a chimeric aadA gene. Proc. Natl. Acad. Sci. USA 90, 913–917 (1993).

    Article  CAS  Google Scholar 

  20. Khan, M.S. & Maliga, P. Fluorescent antibiotic resistance marker for facile identification of transplastomic clones. Nat. Biotechnol. 17, (1999).

  21. Maliga, P. Towards plastid transformation in flowering plants. Trends Biotechnol . 11, 101–107 ( 1993).

    Article  CAS  Google Scholar 

  22. Takeshima, Y., Duguira, M. & Hagiwara, H. A novel expression vector for the cyanobacterium Synechococcus PCC 6301. DNA Res. 1, 181–189 (1996).

    Article  Google Scholar 

  23. Daniell. H. et al. Transient foreign gene expression in chloroplasts of cultured tobacco cells after biolistic delivery of chloroplast vectors. Proc. Natl. Acad. Sci. USA 87, 88–92 (1993).

    Article  Google Scholar 

  24. Daniell, H., Krishnan, M. & McFadden, BF. Transient expression of beta-glucuronidase in different cellular compartments following biolistic delivery of foreign DNA into wheat leaves and calli. Plant Cell Rep. 9, 615 –619 (1991).

    Article  CAS  Google Scholar 

  25. Rippka, R., Waterbury, D.J., Herdman, M. & Stanier, R.Y. Generic assignments, strain histories and properties of pure cultures of cyanobacteria. J. Gen. Microbiol. 111, 1– 61 (1979).

    Google Scholar 

  26. Handler, J.S., Perkins, F.M. & Johnson, J.P. Hormone effects on transport in cultured epithelia with high electrical resistance. Am. J. Physiol. 240 , 103–105 (1981).

    Article  Google Scholar 

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Acknowledgements

We extend our thanks to Dr. Hubert Felle (University of Giessen, Germany), Dr. Winfried Peters (University of Frankfurt, Germany), and Dr. Walter Schröder (Nuclear Research Centre, Juelich, Germany), for helpful technical discussions, Dr. Ursula Richter (University of Giessen, Germany) and the ZBB (University of Giessen) for scanning electron microscope imaging of the pipette tips, Dr. Wolfgang Clauβ (University of Giessen) for Xenopus distal kidney tubule A6 cell cultures, and the Geraberger Thermometer Werke GmbH for a supply of galinstan for the first tests. We also thank Dr. Chris Howe (Department of Biochemistry, University of Cambridge, UK) for supplying P. laminosum. We thank the EU for Grant BIO4 CT972245 as part of the Plasmonet consortium for funding.

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Authors and Affiliations

  1. Institute of General Botany and Plant Physiology, Justus Liebig University Giessen, Senckenbergstrasse 17, Giessen, 35390, Germany

    Michael Knoblauch & Aart J.E. van Bel

  2. Department of Plant Sciences, University of Cambridge, Cambridge, CB2 3EA, Downing Street, UK

    Julian M. Hibberd & John C. Gray

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  1. Michael Knoblauch
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  2. Julian M. Hibberd
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  4. Aart J.E. van Bel
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Correspondence to Michael Knoblauch.

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Knoblauch, M., Hibberd, J., Gray, J. et al. A galinstan expansion femtosyringe for microinjection of eukaryotic organelles and prokaryotes. Nat Biotechnol 17, 906–909 (1999). https://doi.org/10.1038/12902

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