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Spider silk-like proteins derived from transgenic Nicotiana tabacum

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Abstract

The high tensile strength and biocompatibility of spider dragline silk makes it a desirable material in many engineering and tissue regeneration applications. Here, we present the feasibility to produce recombinant proteins in transgenic tobacco Nicotiana tabacum with sequences representing spider silk protein building blocks . Recombinant mini-spidroins contain native N- and C-terminal domains of major ampullate spidroin 1 (rMaSp1) or rMaSp2 flanking an abbreviated number (8, 16 or 32) of consensus repeat domains. Two different expression plasmid vectors were tested and a downstream chitin binding domain and self-cleavable intein were included to facilitate protein purification. We confirmed gene insertion and RNA transcription by PCR and reverse-transcriptase PCR, respectively. Mini-spidroin production was detected by N-terminus specific antibodies. Purification of mini-spidroins was performed through chitin affinity chromatography and subsequent intein activation with reducing reagent. Mini-spidroins, when dialyzed and freeze-dried, formed viscous gelatin-like fluids.

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Acknowledgments

The authors would like to thank Dr. Indu Maiti at the Kentucky Tobacco Research and Development Center for the tobacco expression plasmids and Allison Nelson for technical support. This work was funded by grant 1R15EB007403-01 from the National Institute of Biomedical Imaging and Bioengineering to WRM.

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Author notes

  1. Charlene Gravgaard

    Present address: College of Pharmacy, Medical University of South Carolina, Charleston, SC, 29425, USA

  2. Heather McCartney

    Present address: Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN, 37232, USA

Authors and Affiliations

  1. Department of Genetics and Biochemistry, Clemson University, 130 McGinty Court, 153 Robert F. Poole Agricultural Center, Clemson, SC, 29634, USA

    Congyue Annie Peng, Julia Russo, Charlene Gravgaard, Heather McCartney, William Gaines & William R. Marcotte Jr.

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  1. Congyue Annie Peng
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  2. Julia Russo
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  3. Charlene Gravgaard
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  5. William Gaines
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Correspondence to William R. Marcotte Jr..

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Charlene Gravgaard and Heather McCartney have contributed equally to this work.

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Peng, C.A., Russo, J., Gravgaard, C. et al. Spider silk-like proteins derived from transgenic Nicotiana tabacum . Transgenic Res 25, 517–526 (2016). https://doi.org/10.1007/s11248-016-9949-1

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