The Genome Full-length Cloning and Invasive Cloning Construction of Cu-cumber Mosaic Virus Banana Isolate
Bananas(Musa spp.),an important tropical cash crop.In order to understand the mechanism of cucumber mosaic virus(CMV)infection of bananas and then to prevent and control them in a targeted manner,cucumber mo-saic virus(CMV-Ban)was extracted from infected banana samples,and its triplet genome RNA was cloned and se-quenced.Among them,cmV-Ban RNA1 has a total length of 3 375 bp,encoding the 1a protein;CMV-Ban RNA2 is 3 043 bp in total length and encodes 2a and 2b proteins;CMV-Ban RNA3 is 2 218 bp in total length and encodes 3a kinematic protein and shell(CP)protein.By BLAST comparison,the nucleotide sequence of CMV-Ban RNA1 had the highest homology with CLW2 RNA1 of the Malaysian cucumber isolate(92.1％),the nucleotide sequence of CMV-Ban RNA2 had the highest homology with the Italian pepper isolate(Vir)(91.8％),and the homology of CMV-Ban RNA3 had the highest homology with the domestically reported rapeseed isolate YN RNA3(94.5％).Compared with Vector NTI Advance 11.5,the 20 amino acid sequence homology of CMV-Ban and Taiwan banana isolates reached 99.9％.Phylogenetic tree analysis showed that the isolate CMV-Ban isolated from Hainan bananas infected with cu-cumber mosaic virus belonged to subgroup IB.After that,the CMV-Ban tripenomy genomic RNA was inserted into the plant expression vector pCass4-RZ using In-Fushion seamless cloning,and the invasive vectors RC-CMV-Banl,RCCMV-Ban2,RC-CMV-Ban3 were successfully constructed,and Nicotiana benthamiana was infected by Agro-bacterium osmotic injection technology,and the leaves showed obvious mosaic symptoms after 21 days,indicating that the infected clone RC-CMV-Ban successfully infected Nicotiana benthamiana.The results provide a scientific basis for the subsequent analysis of the infection mechanism and control of banans by CMV-Ban.
Musa spp.Cucumber mosaic virusEvolutionary tree analysisInvasive cloning
NETL
NSTL
万方数据
维普
