李属坏死环斑病毒新疆分离物运动蛋白基因片段的克隆与序列分析

Cloning and sequence analysis of MP gene of Prunus necrotic ringspot virus from almond plants in Xinjiang

为进一步揭示李属坏死环斑病毒（Prunus necrotic ringspot virus,PNRSV）新疆巴旦木分离物的分子特征、遗传变异及其与宿主之间的相互关系,采用RT-PCR方法扩增并克隆了4个PNRSV新疆巴旦木分离物运动蛋白（move protein,MP）基因片段,并进行了测序及序列同源性分析。结果表明,4个新疆巴旦木PNRSV分离物MP基因片段分别为259、258、254、260 bp;其核苷酸和氨基酸序列与已报道的PNRSV分离物的同源性分别为72.7%~91.7%和75.6%~92.9%,表现出明显差异,其中与美国分离物CH9同源性最高,分别达88.8%~91.7%和82.6%~92.9%;而与同属03亚组的苹果花叶病毒（Apple mosaic virus,Ap MV）同源性较低,仅为51.2%~58.1%和52.3%~61.9%;新疆PNRSV各分离株之间MP基因核苷酸序列同源性较高。系统发育树显示,4个新疆巴旦木PNRSV分离物与Ⅰ组代表毒株PV32的核苷酸序列同源性达88.4%~91.3%,并与Ⅰ组分离物聚集成簇,表明PNRSV新疆巴旦木分离物属于引起严重症状的Ⅰ组株系,且Ⅰ组中各分离物之间表现出一定的寄主相关性,而Ⅱ组和Ⅲ组中各分离物之间未表现出明显的寄主相关性。

In order to reveal molecular characteristics,genetic variation of Prunus necrotic ringspot virus（ PNRSV） and the interaction between the virus and its host,the movement protein（ MP） genes of four isolates of four PNRSV were amplified from almond plants in Xinjiang by using the RT-PCR method,sequencing and sequence homology analysis. The sequencing results showed that the lengths of MP gene of four PNRSV-Xinjiang isolates were 259,258,254 and 260 bp,respectively. The similarity analysis of the nucleotide sequences and deduced amino acids of MP genes showed remarkable differences between PNRSV-Xinjiang isolates and those from several different countries,sharing identities of 72. 7% to91. 7% and 75. 6% to 92. 9%,respectively. They shared the highest identity with PNRSV-America isolates CH9,with a homology rate of 88. 8%- 91. 7% and 82. 6%- 92. 9% for nucleotide sequencesand amino acids,respectively,but less than 51. 2%- 58. 1% and 52. 3%- 61. 9% with Apple mosaic virus（ Ap MV） that belongs to the same subgroup of Ilarvirus. High sequence identities of MP genes were shown among Xinjiang isolates. Phylogenetic analysis showed that the four Xinjiang isolates shared the highest homology（ 88. 4%- 91. 3%） with group Ⅰ representative strain PV32. Besides,PNRSVXinjiang isolates clustered together with isolates in groupⅠ. It was verified that PNRSV-Xinjiang isolates belonged to group Ⅰ（ the serious pathotype） and there were more distinct host relationships among groupⅠ isolates analyzed; however,there was no obvious host relationships between various isolates in groupⅡ and group Ⅲ.