The Identification Of PVY Strains On Tobacco In Hunan Province

In recent years, the occurrance of Potato virus Y has expanded across the country, and the degree of its damage has risen. PVY had broken out across some tobacco-growing areas of Hunan province in 2008 and 2009, which caused a serious loss on tobacco and potato in those areas. At present, little was known about the types of PVY strains and their distribution, so it was required urgently to establish rapid and accurate detection methods to figure out the incident of PVY on tobacco in Hunan Province, which could lay the foundation for the effective controlling of the diseases and breeding programs for resistance.Single RT-PCR was established for identifying PVYN, PVYO and PVYNTN through tobacco leaves collected from main tobacco-growing areas in Hunan in 2009 and 2010 which were suspected to infect PVY. By optimizing the annealing temperature and the concentration of primers and Mg2+, a mRT-PCR detecing PVYN, PVYO and PVYNTN simultaneously was erected on the basis of single RT-PCR. This reseach found that the annealing temperature which was determined based on the principle of TdPCR is more suitable for mRT-PCR. A fixed temperature as the annealing temperature can also be used in mRT-PCR, but it is noticeable that the annealing temperature in mRT-PCR should be lower than that in single RT-PCR for about 3℃. If the concentration of primers were not adjusted, the homogeneousness of bands was very poor. The principle of adding primers is to get an increase in the amount of primers for the "weak" loci and a decrease in the amount for the "strong" loci. When the concentration of Mg2+ is optimized, it just require to pay attention to the ratio of concentrations of Mg2+ and dNTP.Collected samples were detected by single RT-PCR and mRT-PCR. The results showed that, PVYN was the major strain of PVY in tobacco-growing areas in Hunan province, and PVYO accoured a bit. PVYNTN was detected firstly on tobacco in Hunan, and PVYO and PVYN:O did not exist. Combined with the analysis of all samples collected in the last two years, it was found that the occurrance of PVY0 and PVYNTN showed an upward trend, although the situation of which in different areas in Hunan was variable. Meanwhile, mixed infections of PVY strains were found on tobacco in Hunan, which mainly belonged to the mixture of PVYN and PVYNTN. The mixture of PVYN and PVYO also existed, but no mixture of PVYO and PVYNTN was detected. The proportion of PVYNTN participating in mixed infection was quit high, which accounted for nearly 50% of total PVYNTN isolates detected.