title: Molecular cloning and in-silico characterization of high temperature stress responsive pAPX gene isolated from heat tolerant Indian wheat cv. Raj 3765.
abstract: BACKGROUND
  Heat stress leads to accelerated production of reactive oxygen species (ROS) which causes a huge amount of oxidative damage to the cellular components of plants. A large number of heat stress related genes as HSPs, catalases, peroxidases are overexpressed at the time of stress. A potent stress responsive gene peroxisomal ascorbate peroxidase (TapAPX) obtained from heat stress (42 °C) responsive subtractive cDNA library from a thermo tolerant wheat cv. Raj3765 at anthesis stage was cloned, characterized and its role was validated under heat stress by proteomics and in-silico studies. In the present study we report the characterization at molecular and in-silico level of peroxisomal TapAPX gene isolated from heat tolerant wheat cultivar of India.
  
  
  RESULTS
  qPCR studies of TapAPX gene displayed up to 203 fold level of expression at 42 °C heat stress exposure. A full length cDNA of 876 bp obtained by RACE deduced a protein of 292 amino acid residues which gives a complete 3D structure of pAPX by homology modeling. TapAPX cDNA was cloned in expression vector pET28 (a+) and the recombinant protein over-expressed in E. coli BL21 showed highest homology with APX protein as deduced by peptide mass fingerprinting.
  
  
  CONCLUSIONS
  TapAPX gene from wheat cv Raj3765 has a distinct role in conferring thermo tolerance to the plants and thus can be used in crop improvement programmes for development of crops tolerant to high temperature.
authors.affiliation: Biotechnology and Climate Change Laboratory, National Research Centre on Plant Biotechnology, Pusa Campu, New Delhi 110012, India. jasdeep_kaur64@yahoo.co.in.
authors.forename: Jasdeep Chatrath
authors.initials: JC
authors.lastname: Padaria
authors.forename: Harinder
authors.initials: H
authors.lastname: Vishwakarma
authors.forename: Koushik
authors.initials: K
authors.lastname: Biswas
authors.forename: Rahul Singh
authors.initials: RS
authors.lastname: Jasrotia
authors.forename: Gyanendra Pratap
authors.initials: GP
authors.lastname: Singh
mesh_terms: D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D060387:Ascorbate Peroxidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods; D018506:Gene Expression Regulation, Plant; D018869:Heat-Shock Response / Q000235:genetics*; D006358:Hot Temperature; D015227:Lipid Peroxidation; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010802:Phylogeny; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011487:Protein Conformation; D040901:Proteomics / Q000379:methods; D015203:Reproducibility of Results; D013329:Structure-Activity Relationship; D014908:Triticum / Q000201:enzymology / Q000235:genetics*
chemical_list: D010940:Plant Proteins; D060387:Ascorbate Peroxidases