1.11.10
EFFICIENCY OF 2'-5'-OLIGOADENYLATES IN ELIMINATION OF PVX, PVS, PVM, PVY, PVF and PLRV FROM TISSUE CULTURE OF POTATO

GI KONOVALOVA, GA YAKOVLEVA and SA BANADYSEV

Belarusian Research Institute for Potato Growing, Samokhvalovichi, Minsk Region, 223013 Belarus

Background and objectives
Inoculation of potato with PVX and PVY activates the 2'-5'-oligoadenylate synthetase [1]. This enzyme polymerizes ATP to a series of 2'-5'-oligoadenylates (2-5A), whose monomers are linked via 2'-5' bonds rather than the usual 3'-5' linkage in the presence of the double-stranded RNA. The application of 2-5A increases the productivity of meristem-derived plants which are free of potato virus X, S, M and F [2]. The aim of this work is the comparative analysis of artificially synthesized 2-5A for elimination of potato virus from tissue culture of affected plants.

Materials and methods
The sprouts of affected tubers of cvs Belorusky ranny, Loshitsky, Orlyonok, Belorusky 3, Lugovskoy and Ramonak were planted on MS medium with addition of 2-5A. In 3-4 weeks, explants of 1-1.5 mm were excised from apical and axillary buds and were passed onto the same medium. Controls: (i) the explants on the medium without 2-5A; (ii) the meristem of 0.1-0.3 mm from sprouts of diseased tubers or plants in vitro which were incubated for 30 days at 37C (heat therapy). Nine tested 2-5A oligomers were synthesized at the Institute of Bioorganic Chemistry (Belarus): numbers 1 and 2 were similar to commercial substances; numbers 7, 8 and 9 were analogues of trimers of 2'-5' oligoadenylic acid that were synthesized on the basis of virazole. In this case, virazole was used as the third control. ELISA was used for testing viruses.

Results and conclusions
The time of regeneration of shoots from explants depended on cultivar, type and concentration of 2-5 A. The rate of regeneration for all varieties and trimers excluding No. 3 was in the range of 47.5-100%, without 2-5A it was 50-82.5%, and for meristem 45-52.5%. Plantlet regeneration was 20-33 days for explants and 60-150 days for meristem. 2-5A of No. 3 inhibited the regeneration rate to 5-12.5%, but the productivity of healthy plants was 80-100%. The amount of free-virus plantlets was equal to zero for control 1 and 38.8-42.8% for control 2. In the presence of virazole the yield of healthy plants was in the range of 36.1-87.5%. The productivity of free-virus plants depended on variety, type and concentration of 2-5A and constituted 95-100% for 2-5A under Nos 7, 8 and 9 at the optimum concentrations. According to the ELISA test, disease level in the 5th progeny of field-grown plants was 5%, constituting 17% as compared to meristem-derived plants by heat therapy method. The absence of heat therapy also reduces the risk of dissemination of PSTV. Verification of virus-free plants in vitro by PCR did not reveal PSTV.

References
1. Babosha AV, Trofimets LN, Ladygina ME, 1990. Doklady Academii Nauk USSR 313, 252-255.
2. Tkachuk ZU, Artemenko VS, Semernikova LI, 1993. Biopolimery Kletka 9, 9-18.