title: Molecular cloning, characterization and differential expression of novel phytocystatin gene during tropospheric ozone stress in maize (Zea mays) leaves.
abstract: In this study, a full-length cDNA encoding a novel phytocystatin gene, designated CC14, was identified in maize leaves. The CC14 gene sequence reported in this study has been deposited in the GenBank database (accession number JF290478). The CC14 gene was cloned into an expression vector pET30 EK/LIC and was then transformed into Escherichia coli strain BL21 (DE3) pLysS to produce a recombinant CC14 protein. The recombinant protein was purified by nickel nitrilotriacetic acid affinity chromatography after induction with 1 mM IPTG. The purified CC14 protein was electrophoresed on SDS-PAGE and a protein 25 kDa in size was observed. Antiprotease activities of the purified recombinant CC14 protein against cysteine proteases and commercially available papain were tested. The results showed that CC14 purified protein suppressed 100% activity of papain and 57-86% plant cysteine protease activity. Moreover, an upregulation of CC14 gene expression was observed after 20 days of ozone stress in maize leaves. Together, these observations concurred to conclude that CC14 gene could potentially be used as a basis for the development of transgenic crops and natural pesticides that resist biotic and abiotic stresses.
authors.affiliation: IBIOS, UMR 7618 Bioemco, Université Paris Est-Créteil, 61, avenue du Général-de-Gaulle, 94010 Créteil cedex, France; Department of Environmental Sciences, COMSATS Institute of Information Technology, 22060 Abbottabad, Pakistan. Electronic address: drrafiq@ciit.net.pk.
authors.forename: Rafiq
authors.initials: R
authors.lastname: Ahmad
authors.affiliation: IBIOS, UMR 7618 Bioemco, Université Paris Est-Créteil, 61, avenue du Général-de-Gaulle, 94010 Créteil cedex, France.
authors.forename: Yasmine
authors.initials: Y
authors.lastname: Zuily-Fodil
authors.affiliation: IBIOS, UMR 7618 Bioemco, Université Paris Est-Créteil, 61, avenue du Général-de-Gaulle, 94010 Créteil cedex, France.
authors.forename: Chantal
authors.initials: C
authors.lastname: Passaquet
authors.affiliation: Department of Environmental Sciences, COMSATS Institute of Information Technology, 22060 Abbottabad, Pakistan.
authors.forename: Sabaz
authors.initials: S
authors.lastname: Ali Khan
authors.affiliation: IBIOS, UMR 7618 Bioemco, Université Paris Est-Créteil, 61, avenue du Général-de-Gaulle, 94010 Créteil cedex, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Repellin
mesh_terms: D003001:Cloning, Molecular; D015891:Cystatins / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D018506:Gene Expression Regulation, Plant / Q000235:genetics; D010126:Ozone; D018515:Plant Leaves; D010940:Plant Proteins / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics*; D015854:Up-Regulation / Q000235:genetics; D003313:Zea mays / Q000235:genetics*
chemical_list: D015891:Cystatins; D018076:DNA, Complementary; D010940:Plant Proteins; D011994:Recombinant Proteins; D010126:Ozone
keywords: CC14; Enzymatic characterization; Gene expression; Maize cystatin; Tropospheric ozone