| The potato is the world's fourth largest food crops, it is an annual solanaceae solanum crops and has the very high nutritional value and use value. Potato is vulnerable to viruses, it has been reported that more than 35 kinds of virus can infect potato, six kinds of virus do most harm to potato industry in sichuan province,they are. potato virus X (PVX), potato virus Y (PVY), potato virus S (PVS), potato A virus (PVA), potato leaf curl virus (PLRV), potato M virus (PVM).Research has shown that 6 kinds of virus can not only infect potatoes by itself, but also can cause compound infection seriously affecting the yield and quality of potatoes.Sichuan did not investigate the distribution of the six main potato viruses since 2006.To understand the distribution of the 6 kinds of virus in sichuan province, in this text we use the methord of serology double antibody sandwich enzyme linked immunosorbent assay to text 1003 potato samples from the field of different altitude like below 500 m,500-1000 m and beyond 500 m..test results showed that six main potato virus in sichuan province all cause different degrees of disease, the incidence of PVA was 10.87%, the incidence of PVM is 5.68%, the incidence of PLRV was 5.38%, the incidence of PVY was 19.44%, the incidence of PVS is 26.44%, the incidence of PVX is 18.74%. In 2006, sichuan province carried out a survey for the distribution of potato virus, the results showed that the incidence of disease of PVA, PVM, PLRV, PVY, PVS and PVX are 169%,9.58%,3.58%,9.58%,61.77% and 17.74%, compared with the results of the survey in 2015,the incidence of PVS virus reduced by 35%, the incidence of PVA reduced by 5%, the incidence of PVY increased by nearly 12%, the incidence of the remaining three viruses haven't changed much, in general, except for PVY. the rest of the five kinds of virus infection are to some extent under control, it has much to do with the planting of potato virus-free seed in recent years.Incidence of different elevation is different, in general. Potato in elevation under 500 m and 500-1000 have a higher incidence,1000 m above have a relatively low incedence; Each virus has different incidence different altitude, PVA and PVY have higher incidence, at an altitude of 500 m below. PLRV, PVM have higher incidence at an altitude of 500-1000.of PVS at an altitude of 500-1000-m area, higher incidence of PVX at an altitude of 1000 m above the region; PVM and PLRV viruses have lowest incidence rates in different altitude and different season,it shows different elevations have a certain impact of six main potato virus, but the season do not cause big infection.From 2014 to 2015, we test 4793 samples in 25 companies in Sichuan usingDAS-ELISA..Results show that the PVS has the highest detection rate at 5.3%, followed by PVM and PVY, detection rate of which are 2.09% and 2.04%. detection rate of PVA, PVX and PLRV under 1%.The production of virus-free potato in Sichuan mainly use the basic seedlings, the original species, native species and production Detoxification of potato production 4 system. The qualified rate of basic seedlings is 96.75%,of witch the original seed is 99.53%, seed is 83.74%, the production is 52.24%.Although the basic seedlings, the original species have a relatively high qualified rate but has not reached the requirements of the 100% purity, therefore, to improve the quality of virus free potato seed from the source, timely prevention should be provided in production.to reduce de occurrence of virus.At present, DAS-ELISA is the main method to detect potato virus, but because of its the high cost, lack of sensitivity, in recent years, molecular biology method has been widely used in the detection of potato virus. This experimental establish the system who can detect PVA PVM, PLRV, PVY, PVS at the same time. witch is a multiple rt-pcr system. We design specific primer according to (CP) gene sequence of 6 kinds of virus published in GenBank, and optimized some main influence factors during PCR,such as Taq enzyme concentration, concentration of dNTPs, annealing temperature, cycles times, results show that 6 kinds of virus can be detected at the same time by some clear and bright stripes is in the system of 25 ul with Taq enzyme in the final concentration of 0.03 U/ul, the final concentration of 0.16 dNTPs mmol/L, annealing temperature for 54? during response procedures and cycles of 35. After sequencing, we BLAST the sequence of 6 kinds of virus in NCBI, and find the homology with reported corresponding virus is above 97%,proving that 6 kinds of virus was amplified accurately; Sensitivity test results showed that the method can detect the minimum concentration of 10-3 amount of organization; Using the multiple rt-pcr reaction system to test to 16 samples of field. The multiple rt-pcr reaction system can detect 6 kinds of potato virus infecting by single or compound rapidly, accurately, sensitively. |
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