S14 - Session P5 - Development of molecular diagnosis of Chinese yam necrotic mosaic virus infecting Dioscorea spp
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Authors: Mei-Ju Lin *, Ting-Chin Deng, Chia-Shin Tsai, Yi-Ting Lin
Yam consistently propagated with tubers or bulbils, therefore some viral diseases had occurred in the field after the long-term asexual reproduction. Plants of Taiwanese Yam ( Dioscorea japonica var. pseudojaponica ) with mosaic symptoms grown from bulbils were obtained. The RT-PCR (reverse transcription- polymerase chain reaction) was performed with a primer pair CYV1665P and CYV1906M. A 251 bp nucleotide fragment was amplified and sequenced, and the nucleotide was showed 99.2% identical to Chinese yam necrotic mosaic virus (CYNMV). The RT-PCR primers were further designed and performed step by step to sequence the whole genome, including the 864 bp capsid protein (CP) gene. The analysis of the CP gene sequence and compared with other viruses belonging to the same genus of Macluravirus revealed that the virus isolate (CYNMV-TY) infecting Taiwanese yam was closely related to Korean isolate of CYNMV-AD and BRI (Acc No. KU641566, KX352243). They shared the 97.9, 98.6% nucleotide identity and 99.7, 99.0% amino acid similarity of CP gene. As comparing with Yam chlorotic mosaic virus (YCMV isolate Acc No. KT724961), the identity of nucleotide sequence of Taiwan isolate CYNMV-TY) was only 71.8% and the amino acid similarity was 69.8%. And comparing with Yam chlorotic necrosis virus (YCNV isolate Acc No. MG755240), the identity of nucleotide sequence of Taiwan isolate CYNMV-TY) was only 64.9% and the amino acid similarity was 56.9%. For identification of Chinese yam necrotic mosaic virus with other two Macluravirus of yam, Yam chlorotic mosaic virus and Yam chlorotic necrosis virus. The specific primers CYNMV-1056u (5'-TATCGCCGTCCCTCCTGTCTAAA-3') and CYNMV-1290d (5'-TTGCGGCGGCTCTCCTCTAT-3') were designed within the conserved regions of HC-Pro. Compare to primers CYV1665P and CYV1906M, the specificity and sensitivity CYNMV-1056u and CYNMV-1290d were excellent for diagnosis of CYNMV. In the future, this detection technology can be used for the screening of healthy seedlings of yam.