title: Expression of the truncated tissue plasminogen activator (K2S) gene in tobacco chloroplast.
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.
authors.affiliation: Department of Plant Breeding & Biotechnology, Faculty of Agriculture, Tarbiat Modares University, P.O. Box 14115-336, Tehran, Iran, Maryam_abdoly@yahoo.com.
authors.forename: Maryam
authors.initials: M
authors.lastname: Abdoli-Nasab
authors.forename: Mokhtar
authors.initials: M
authors.lastname: Jalali-Javaran
authors.forename: Rosa M
authors.initials: RM
authors.lastname: Cusidó
authors.forename: Javier
authors.initials: J
authors.lastname: Palazón
authors.forename: Amin
authors.initials: A
authors.lastname: Baghizadeh
authors.forename: Houshang
authors.initials: H
authors.lastname: Alizadeh
mesh_terms: D015139:Blotting, Southern; D015153:Blotting, Western; D002736:Chloroplasts / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005780:Gelatin / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D012038:Regeneration; D010959:Tissue Plasminogen Activator / Q000235:genetics* / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
chemical_list: D005780:Gelatin; D010959:Tissue Plasminogen Activator