The GPRIME package: computer programs for identifying the best regions of aligned genes to target in nucleic acid hybridisation-based diagnostic tests, and their use with plant viruses.

The GPRIME (Group PRIMEr design) programs examine aligned sets of gene sequences to discover homologous regions to be targeted in diagnostic tests. The core program moves a 'window' over the aligned sequences and calculates, at each window position, a 'redundancy value', namely the number of sequences that would represent all permutations of the variable sequence positions within that window. Regions with minimal redundancy values may then be targeted in diagnostic tests based on oligonucleotide hybridisation. The likely specificity of tests targeting such regions can be assessed by searching the international databases with those regions using FASTA. The GPRIME programs, which include programs for designing primers to distinguish between two sub-sets of a group of aligned sequences, can be obtained from http://life.anu.edu.au/software.html. We have used GPRIME to design redundant primers for RT-PCR tests to detect all potexviruses and tobamoviruses, and then used these, together with a previously reported pair of primers for the Potyviridae, to screen some Australian orchid collections. Two orchid viruses previously reported from Australia were found; cymbidium mosaic potexvirus was common, but odontoglossum ringspot tobamovirus was not. In addition the recently described ceratobium mosaic potyvirus was found to be common, and three other novel potyviruses were also found.