title: Sap-direct RT-PCR for the rapid detection of coleus blumei viroids of the genus Coleviroid from natural host plants.
abstract: A simple and fast sap-direct RT-PCR (reverse transcription-polymerase chain reaction) for the rapid detection of 3 viroids of the genus Coleviroid is presented. The templates for cDNA synthesis were obtained directly from the sap of coleus using a pipettor, a common tool in molecular biology laboratories, and 3 coleus blumei viroids (CbVds) were detected simultaneously using a pair of universal primers designed according to sequences in the central conserved region (CCR) of CbVds. RT-PCR results demonstrated that CbVd-1, CbVd-5, and CbVd-6 can be detected accurately in viroid-infected plants but not in viroid-free plants. The results of RT-PCR, dot-blot, sequencing, and batch-detection revealed that this method can be used to identify CbVds rapidly. The method also reduces cross-contamination among different samples to a minimum. It is considered that this rapid and simple technique is an effective method for the identification and cloning of CbVds.
authors: Institute of Plant Virology, Fujian Agriculture and Forestry University, Fuzhou 350002, Fujian Province, China.
authors: Dongmei
authors: D
authors: Jiang
authors: Zujian
authors: Z
authors: Wu
authors: Lianhui
authors: L
authors: Xie
authors: Teruo
authors: T
authors: Sano
authors: Shifang
authors: S
authors: Li
mesh_terms: D031330:Coleus / Q000821:virology*; D017931:DNA Primers / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014772:Viroids / Q000235:genetics / Q000302:isolation & purification*; D014773:Virology / Q000379:methods*
chemical_list: D017931:DNA Primers; D012367:RNA, Viral