Simultaneous detection of one RNA and one DNA virus from naturally infected citrus plants using duplex PCR technique.
Citrus tristeza clostero virus (CTV), an aphid-transmitted RNA virus having a genome size of about 19.3 kb, singly or as a mixed infection with Citrus mosaic badna virus (CMBV), a non-enveloped bacilliform DNA virus having genome of 7.5 kb, plays a significant role in causing citrus decline, particularly in sweet orange in India. Rapid detection techniques are important in the prevention of spread of these two diseases in field conditions. Since CMBV is weakly immunogenic, sero-diagnosis is not the preferred diagnostic method. Similarly, for detection of CTV though serological techniques like ELISA are being widely used, production of polyclonal antibodies to various isolates of CTV is often limited by various factors, namely low yields of the virus in plant tissues, uneven distribution, difficulties in production of sufficient quantities of infected tissue, contamination by host proteins in purified preparation, international quarantine, etc. As an alternative, a rapid and reliable PCR based detection protocol has been standardized. Sets of primers were designed based on the respective virus isolate sequence data available in GenBank, to obtain anticipated products of calculated size.