In vitro production of Indian citrus ringspot virus (ICRSV) free kinnow plants employing phytotherapy coupled with shoot tip grafting

Abstract

This paper reports the elimination of Indian citrus ringspot virus (ICRSV) from “kinnow” (Citrus nobilis Lour × Citrus deliciosa Tenora) employing phytotherapy coupled with shoot tip grafting under in vitro conditions. Nodal segments from infected mother plant (indexed by indirect enzyme-linked immunosorbent assay [ELISA] and reverse transcriptase PCR [RT-PCR]) were cultured on Murashige and Skoog medium containing 2iP (1 mg/l or 4.9 μM) and malt extract (800 mg/l) along with different concentrations of aqueous extracts from leaves of Azadirachta indica (Neem), Sorghum vulgare (Jowar), and roots of Boerhaavia diffusa (Punarnava). Shoot tips were excised from the nodal sprouts and grafted on to rough lemon (Citrus jambhiri) under aseptic conditions. Maximum effect (50% virus elimination) was seen for aqueous leaf extracts of A. indica followed by B. diffusa root extract (42.86%) and S. vulgare leaf extract (31.58%). Plants/plantlets were considered virus-free only when showing negative reactions by both indirect ELISA and RT-PCR.