Progressive Agriculture
  • Year: 2018
  • Volume: 18
  • Issue: 1

Development of transgene construct using coat protein gene of papaya ringspot virus and its validation

  • Author:
  • Ashwini Kumar1, YB Basavaraj2, Pushpendra Kumar1, Manoj Kumar Yadav1, Prashant Mishra3, Shiv Kumar Singh4
  • Total Page Count: 6
  • Page Number: 92 to 97

1Deptt. of Agricultural Biotechnology, Sardar Vallabhbhai Patel University of Agriculture and Technology, Meerut, 250110, India

2Division of Plant Pathology, Indian Agriculture Research Institute, New Delhi, 110012, India

3Department of Plant pathology, Sardar Vallabhbhai Patel University of Agriculture and Technology, Meerut, 250110, India

4Department of Genetics and Plant Breeding, Sardar Vallabhbhai Patel University of Agriculture and Technology, Meerut, 250110, India

Online published on 10 April, 2018.

Abstract

Papaya ringspot virus (PRSV), continues to be a major threat for the production of papaya as well as cucurbits in India. The virions of PRSV possess a single stranded translatable RNA genome of 10317–10349 nt encoding a polyprotein of 3341 to 3347 aa. Of 10 viral proteins derived from polyprotein, coat protein (CP) has been studied extensively. The coat protein mediated resistance (CPMR) strategy has shown promise in Hawaii (USA) in managing PRSV and thus explored the strategy in present study. Developed Full CP-based construct (pCAMBIA1302: FullCP) along with GFP using isolates as a transgene source based sequence homology analysis The construct was validated for physical integrity (through PCR and restriction digestion analysis) as well as for biological functionality (through transient expression of transgene proteins i.e. CP & GFP by agro-infiltration method in the experimental host plants). The construct could express the GFP at inter and intra cellular level as visualized under confocal microscopy. Further, the construct could express the coat protein in the infiltrated leaf tissues as analyzed through DAC-ELISA based serological assay using PAbs to PRSV-CP. This construct found structurally as well as functionally active under plant system and can be further modified into transgene construct to be used to raise the transgenic papaya lines to fight against PRSV infections.

Keywords

PRSV, Construct