ToCV和TYLCV复合侵染番茄后部分病毒基因序列分析及实时荧光定量PCR分析
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摘要
番茄褪绿病毒病(ToCV)和番茄黄化曲叶病毒(TYLCV)存在复合侵染的现象,其发病率逐年递增,成为一种新的威胁番茄产业的病毒病害。为了研究复合侵染样品中2种病毒的互作和基因变异情况,对天津番茄产区发生的TYLCV全基因组和ToCV的CP基因和HSP基因进行了克隆和序列分析。结果表明,与单独侵染样品进行比对,TYLCV的基因序列在复合侵染的样品中共有6处碱基发生了变异,分别为73位A→C、92位A→G、347位C→T、1 209位C→T、1 618位A→G、2 107位A→T,除73位和92位的变异未在编码区,其余的变异碱基均在ORF框内,其中1 209位为同义突变,其他均发生了错义突变。ToCV的CP基因有1处同义突变,ToCV的HSP基因共有4处碱基变异,其中826位由T→C和1 166位由G→A均为同义突变,1 395位和1 630位均由A→G,为错义突变。利用实时荧光定量PCR技术对单独侵染和复合侵染的番茄样品中TYLCV和ToCV病毒积累量进行分析。结果表明,在田间单独侵染和复合侵染的样品中, TYLCV的拷贝数差异不显著,ToCV的拷贝数在单独侵染和复合侵染的样品中差异也不显著。在复合侵染的样品中,TYLCV的拷贝数约为ToCV的262～436倍。
There exists the compound infection of Tomato chlorosis virus(ToCV) and Tomato yellow leaf curl virus(TYLCV), and the incidence of the two diseases presents a tread of increase in recent years, which becomes a new threat to tomato farming. In order to study the interaction and variation of the two virus genes after single and mixed infection, the molecular identification and sequence analysis of the CP gene and the HSP gene of ToCV and the gene of TYLCV occurred in Tianjin were carried out. The results showed that, compared with the single infection samples, 6 base mutations of TYLCV were found in mixed infection samples, i.e. 73 rd A→C, 92 nd A→G, 347 th C→T, 1 209 th C→T, 1 618 th A→G and 2 107 th A→T, among which the 73 rd and 92 nd mutations were not in the coding region and the other four were located in the ORF box with 3 missense mutations and one samesense mutation(1 209 th). There was also one samesense mutation in the CP gene of ToCV. There were 4 base mutations in HSP gene of ToCV, among which the 826 th T→C and 1 166 th G→A were samesense mutations, while the 1 395 th A→G and 1 630 th A→G were missense mutations. The accumulation amount of TYLCV and ToCV in tomato samples after single infection and mixed infection were analyzed by RT-qPCR. The results showed that the accumulations of TYLCV and ToCV had no significant difference between single infection and mixed infection. In the mixed infection samples, the copy of TYLCV was about 262-436 times that of ToCV.
There exists the compound infection of Tomato chlorosis virus(ToCV) and Tomato yellow leaf curl virus(TYLCV), and the incidence of the two diseases presents a tread of increase in recent years, which becomes a new threat to tomato farming. In order to study the interaction and variation of the two virus genes after single and mixed infection, the molecular identification and sequence analysis of the CP gene and the HSP gene of ToCV and the gene of TYLCV occurred in Tianjin were carried out. The results showed that, compared with the single infection samples, 6 base mutations of TYLCV were found in mixed infection samples, i.e. 73 rd A→C, 92 nd A→G, 347 th C→T, 1 209 th C→T, 1 618 th A→G and 2 107 th A→T, among which the 73 rd and 92 nd mutations were not in the coding region and the other four were located in the ORF box with 3 missense mutations and one samesense mutation(1 209 th). There was also one samesense mutation in the CP gene of ToCV. There were 4 base mutations in HSP gene of ToCV, among which the 826 th T→C and 1 166 th G→A were samesense mutations, while the 1 395 th A→G and 1 630 th A→G were missense mutations. The accumulation amount of TYLCV and ToCV in tomato samples after single infection and mixed infection were analyzed by RT-qPCR. The results showed that the accumulations of TYLCV and ToCV had no significant difference between single infection and mixed infection. In the mixed infection samples, the copy of TYLCV was about 262-436 times that of ToCV.
引文
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[13] 曹骞,李晶,买热木古丽·克依木,王惠卿,李国志,马德英.新疆地区烟粉虱生物型的区域分布及其携带的番茄黄化曲叶病毒检测[J].昆虫学报,2013,56(6):652-664.Cao Q,Li J,Mairemuguli K Y M,Wang H J,Li G Z,Ma D Y.Biotype distribution and infection status of tomato yellow leaf curl virus(TYLCV) in Bemisia tabaci(Hemiptera:Aleyrodidae) in Xinjiang,northwestern China[J].Acta Entomalogica Sinica,2013,56(6):652-664.
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