title: Biotechnological production of inducible defense-related proteins in edible radish (raphanus sativus) found in Nepal.
abstract: BACKGROUND
  Fungal infection in plant leads to use of many hazardous antifungal chemicals. Alternative to these chemicals, defense related antifungal proteins can be used in case of fungal diseases.
  
  
  OBJECTIVE
  An experiment was done in two varieties of edible radish (Raphanus sativus var. Pyuthane Raato and Raphanus sativus var. all season) with aims to produce defense protein within the plant, to identify and perform molecular characterization of those antifungal proteins. The next aim was to compare the antifungal property of those proteins with commercially available synthetic pesticides.
  
  
  METHODS
  Both varieties of radish were infected with fungi (Alternaria alternata and Fusarium oxysporum). Protein samples were isolated from leaves following the standard protocol as described for β-glucuronidase (GUS) assay and were run along with the standard protein marker of 10-250kDa in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to identify and molecularly characterize them.
  
  
  RESULTS
  An additional band in the range of 37-50kDa was observed in the fungal infected samples, which was not seen on uninfected samples. The antifungal assay was carried out for every sample in 96 wells microtitre plate. The extracted protein samples from fungal inoculated plants showed the significant inhibition of fungal growth compared to other samples. On the basis of molecular weight and their antifungal properties, the protein samples from the fungal infected plant were found to be PR2 (Glucanase) and PR3 (Chitinase).
  
  
  CONCLUSIONS
  Defense related proteins were successfully produced in two varieties of radish found in Nepal. The use of such biologically produced proteins may reduce the use of biologically harmful synthetic pesticides.
authors.affiliation: Student, Department of Biotechnology, Himalayan Whitehouse College of Science and Engineering , Lalitpur, Nepal .
authors.forename: Praval
authors.initials: P
authors.lastname: Khanal
authors.affiliation: Student, Department of Biotechnology, Himalayan Whitehouse College of Science and Engineering , Lalitpur, Nepal .
authors.forename: Anil
authors.initials: A
authors.lastname: Karmacharya
authors.affiliation: Lecturer, Department of Biotechnology, Himalayan Whitehouse College of Science and Engineering , Lalitpur, Nepal .
authors.forename: Shishir
authors.initials: S
authors.lastname: Sharma
authors.affiliation: Senior Demonstrator, Department of Biochemistry, B.P. Koirala Institute of Health and Sciences , Dharan, Nepal .
authors.forename: Ashwini K
authors.initials: AK
authors.lastname: Nepal
authors.affiliation: Senior Scientist, National Academy of Science and Technology , Lalitpur, Nepal .
authors.forename: Kanti
authors.initials: K
authors.lastname: Shrestha
keywords: Antifungal protein; Chitinase; Glucanase; Molecular characterization; Raphanus sativus