A Reexamination of the Effectiveness of Ribavirin on Eradication of Viruses in Potato Plantlets in vitro Using ELISA and Quantitative RT-PCR
A Reexamination of the Effectiveness of Ribavirin on Eradication of Viruses in Potato Plantlets in vitro Using ELISA and Quantitative RT-PCR
复制标题
DOI：
10.1007/s12230-013-9350-z
复制
发表时间：
2014-06
影响因子：
1.5
通讯作者：
Lijie Yang;Bihua Nie;Jun Liu;Botao Song
中科院分区：
文献类型：
--
作者：
Lijie Yang;Bihua Nie;Jun Liu;Botao Song
将PVA、PLRV、PVS、PVX、PVY单独感染和PVM、PVS、PVY混合感染的马铃薯试管苗接种在含有不同浓度利巴韦林的MS培养基上。研究了病毒唑对试管苗生长和脱毒效果的影响。结果表明，随着利巴韦林浓度的增加，株高和鲜重明显下降，当浓度达到200 mg/L时，大部分试管苗不能成活。ELISA检测结果表明，利巴韦林对PVA、PVM、PVS和PVX的抑制效果优于PVY和PLRV，且经2 ~ 3次继代培养后，用75 ~ 150 mg/L的利巴韦林进行继代培养，可获得100%的健康苗。而用75 ~ 150 mg/L病毒唑继代3次后，只有33 ~ 66%的PVY和PLRV感染苗脱毒。定量RT-PCR（qPCR）结果表明，病毒唑能明显降低试管苗中的病毒含量。除病毒唑继代3次后检测到PLRV阳性外，在繁殖的第一或末期从感染的砧木中获得健康的幼苗，在根除后阶段未检测到病毒。SSR分析表明，利巴韦林处理引起的遗传变异在对照和处理植株之间没有明显差异。以上结果证明了病毒唑离体处理是一种有效的脱毒方法。
The potato plantlets singly infected by PVA, PLRV, PVS, PVX and PVY and mix-infected by PVM, PVS and PVY were cultured on MS medium with different concentration of ribavirin. The effects of ribavirin on growth of the plantlets and efficiency of virus elimination were investigated. Results showed that the plant height and fresh weight obviously decreased with increase of ribavirin concentration from 0 mg/L to 150 mg/L, and most of the plantlets could not survive when the concentration reached 200 mg/L. According to the ELISA tests, ribavirin was more efficient for eradicating PVA, PVM, PVS and PVX than PVY and PLRV, and healthy plantlets could be obtained with high frequency (up to 100 %) by culturing with 75 ~ 150 mg/L ribavirin after 2 ~ 3 subcultures. Whereas, only 33 ~ 66 % PVY and PLRV infected plantlets were found to be virus-free after 3 subcultures with 75 ~ 150 mg/L ribavirin. The results of quantitative RT-PCR (qPCR) indicated that ribavirin could obviously reduce virus content in the plantlets. Except PLRV was detected positive after 3 subcultures with ribavirin, the healthy seedlings were obtained from infected stocks at the first or end of propagation and no viruses could be detected at the post-eradication stage. No apparent difference of genetic variation resulted from ribavirin treatment was found by SSR analysis between the control and the treated plantlets. All of these results above proved that ribavirin treatment in vitro was an effective method to eliminate viruses in the propagation of potato.