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Expression of the truncated tissue plasminogen activator (K2S) gene in tobacco chloroplast

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Abstract

As because the plant plastid genome is highly polyploid, the transformation of chloroplasts permits the introduction of thousands of copies of foreign genes per plant cell and generates extraordinarily high levels of recombinant protein. Human tissue-type plasminogen activator is one of the most important pharmaceutical proteins involved in the breakdown of blood clots in brain and heart blood vessels. We report the introduction and expression of the truncated human tissue plasminogen activator (K2S) gene in tobacco chloroplasts. The K2S-containing vector pKCZK2S was successfully transferred to tobacco plastomes using the biolistic delivery procedure. Transplastomic plants were selected on RMOP medium containing spectinomycin (500 mg/l). In order to achieve homoplasmy, several rounds of selection and regeneration were performed. The presence, site-specific integration, homoplasmy, expression and activity assay of the transgene were confirmed in the transplastomic plants by PCR, Southern-blot, RT-PCR, SDS-PAGE, ELISA, Dot-blot, Western-blot and zymography analysis. Our results show that the tissue plasminogen activator (K2S form) protein to be expressed in tobacco chloroplasts in active form.

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Authors and Affiliations

  1. Department of Plant Breeding & Biotechnology, Faculty of Agriculture, Tarbiat Modares University, P.O. Box 14115-336, Tehran, Iran

    Maryam Abdoli-Nasab & Mokhtar Jalali-Javaran

  2. Department of Plant Physiology, Faculty of Pharmacy, University of Barcelona, Av. Joan, XXIII, S/n 08028, Barcelona, Spain

    Rosa M. Cusidó & Javier Palazón

  3. Department of Biotechnology, Institute of Science, High Technology & Environmental Sciences, Graduate University of Advanced Technology, P.O. Box 76315-117, Kerman, Iran

    Amin Baghizadeh

  4. Department of Agronomy & Plant Breeding, College of Agriculture & Natural Resources, University of Tehran, P.O. Box 4111, Karaj, Iran

    Houshang Alizadeh

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  1. Maryam Abdoli-Nasab
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  2. Mokhtar Jalali-Javaran
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  3. Rosa M. Cusidó
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  6. Houshang Alizadeh
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Correspondence to Mokhtar Jalali-Javaran.

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Abdoli-Nasab, M., Jalali-Javaran, M., Cusidó, R.M. et al. Expression of the truncated tissue plasminogen activator (K2S) gene in tobacco chloroplast. Mol Biol Rep 40, 5749–5758 (2013). https://doi.org/10.1007/s11033-013-2678-0

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  • DOI: https://doi.org/10.1007/s11033-013-2678-0

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