Articles
STUDY AND CLONING OF A TAIWAN ISOLATE OF HIBISCUS CHLOROTIC RINGSPOT VIRUS
Article number
722_37
Pages
299 – 304
Language
English
Abstract
Hibiscus rosa-sinensis (hibiscus) is grown as a popular hedge or potted flower in Taiwan.
Chlorotic ringspots or spots are frequently observed on the leaves of hibiscus plants.
After using mechanical inoculation, chlorotic local lesions appeared on the leaves of Chenopodium quinoa. An isolate of a virus was obtained via three successive single lesion isolation in C. quinoa. In host range test, H. cannabinus (kenaf), H. rosa-sinensis, and H. sabdariffa (roselle) were infected systemically by the virus.
The partially purified virions were isomeric and approximately 28 nm in diameter by TEM. Using double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), these partially purified virions, as well as symptomatic plants, tested positive using a polyclonal antibody of Hibiscus chlorotic ringspot virus (HCRSV) (Agdia Inc., Elkhart, IN, USA). From the results of host range, TEM and ELISA tests, the virus isolate is proved to be a Taiwan isolate of HCRSV, and thus named HCRSV-TW. The complete genome of HCRSV-TW was cloned and sequenced, and it has 3910 nts containing seven ORFs.
After sequence comparison of the Taiwan and Singapore isolates of HCRSV, the most interesting finding is that the nucleic acid sequence identity of p23 is 84%, but the amino acid sequence identity is only 67%. For other ORFs, percentage identities at the amino acid level range from 83% (p28) to 94% (CP) between two isolates.
In addition, two cDNA clones of HCRSV-TW were constructed with a T7 promoter flanking to the 5 end.
Uncapped HCRSV RNAs transcribed in vitro from the cDNA clones were infectious in protoplasts and plants.
Chlorotic ringspots or spots are frequently observed on the leaves of hibiscus plants.
After using mechanical inoculation, chlorotic local lesions appeared on the leaves of Chenopodium quinoa. An isolate of a virus was obtained via three successive single lesion isolation in C. quinoa. In host range test, H. cannabinus (kenaf), H. rosa-sinensis, and H. sabdariffa (roselle) were infected systemically by the virus.
The partially purified virions were isomeric and approximately 28 nm in diameter by TEM. Using double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), these partially purified virions, as well as symptomatic plants, tested positive using a polyclonal antibody of Hibiscus chlorotic ringspot virus (HCRSV) (Agdia Inc., Elkhart, IN, USA). From the results of host range, TEM and ELISA tests, the virus isolate is proved to be a Taiwan isolate of HCRSV, and thus named HCRSV-TW. The complete genome of HCRSV-TW was cloned and sequenced, and it has 3910 nts containing seven ORFs.
After sequence comparison of the Taiwan and Singapore isolates of HCRSV, the most interesting finding is that the nucleic acid sequence identity of p23 is 84%, but the amino acid sequence identity is only 67%. For other ORFs, percentage identities at the amino acid level range from 83% (p28) to 94% (CP) between two isolates.
In addition, two cDNA clones of HCRSV-TW were constructed with a T7 promoter flanking to the 5 end.
Uncapped HCRSV RNAs transcribed in vitro from the cDNA clones were infectious in protoplasts and plants.
Authors
S.H. Li, Y.C. Chang
Keywords
HCRSV, carmovirus, Hibiscus rosa-sinensis, Hibiscus cannabinus, Hibiscus sabdariffa
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