Shaanxi Tobacco Research Institute

Fengcheng, China

Shaanxi Tobacco Research Institute

Fengcheng, China
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Zhao L.,Northwestern College | Cheng J.,Shaanxi Tobacco Research Institute | Hao X.,Northwestern College | Tian X.,Yangling Vocational and Technical College | Wu Y.,Northwestern College
Archives of Virology | Year: 2012

Tobacco viruses may cause a wide range of diseases that heavily reduce tobacco quality and yield worldwide. In order to detect viral diseases in tobacco fields, a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) method was established. Nucleotide amplification could be observed clearly after adding SYBR Green I, within 60 min under isothermal conditions, at 63-65 °C with a set of primers targeting the viral coat protein (CP) genes of tobacco viruses including cucumber mosaic virus (CMV), potato virus Y (PVY), tobacco etch virus (TEV), tobacco mosaic virus (TMV) and tobacco vein banding mosaic virus (TVBMV). This method has high specificity and sensitivity. The sensitivity of the RT-LAMP was 10 to 100 times higher than that of the conventional RT-PCR method. The RT-LAMP assay was proven reliable for virus diagnosis of tobacco samples from the field. © 2012 Springer-Verlag.

Chen W.,Northwest University, China | Dai J.,Northwest University, China | Zhang H.,Northwest University, China | Jiao H.,Northwest University, China | And 2 more authors.
Turkish Journal of Agriculture and Forestry | Year: 2014

Irrigation water can be polluted by the tobacco etch virus (TEV), which causes serious economic loss in tobacco. However, it is difficult to monitor the sanitary status of irrigation water because TEV presents at extremely low concentrations. This study designed a procedure for concentrating TEV from a standard water sample using polyethylene glycol and detecting it using SYBR Green-based quantitative real-time polymerase chain reaction (qRT-PCR). The concentration factors were optimized through orthogonal tests and the highest recovery efficiency of the TEV genome from a standard water sample was 92.05%. The sensitivity was evaluated using nine 10-fold dilution series of TEV plasmids, and the detection limit of the qRT-PCR was about 10 viral copies/μL. In the infectivity test, TEV was first detected in the roots of NC89 at 7 days after treatment and in the upper leaves at 14 days after treatment. Field diagnosis results showed that 56 of the 180 samples tested positive for TEV, which indicated that this method may be suitable for concentrating and detecting TEV in irrigation water. © TÜBİTAK.

Shan H.,Northwest University, China | Zhao M.,Yangtze University | Chen D.,Qingzhou Tobacco Research Institute of China National Tobacco Crop | Cheng J.,Shaanxi Tobacco Research Institute | And 4 more authors.
Crop Protection | Year: 2013

Strain BC79, isolated from primeval forest soil in Qinling, Mountains, China, was identified as Bacillus methylotrophicus based on morphological, biochemical, physiological and chemotaxonomic analyses as well as phylogenetic 16S rDNA sequencing data. This strain was able to suppress mycelial growth and conidial germination of numerous plant pathogenic fungi in dual cultures on solid media. For exploring potential biocontrol activity, we assessed fermentation conditions for studying B. meth1ylotrophicus BC79. The active substance of BC79, phenaminomethylacetic acid, was extracted by TLC and HPLC, and identified as the strongest inhibitory substance described in B. methylotrophicus. Experiments in a greenhouse showed that application of BC79 culture filtrates 24 h before inoculation of Magnaporthe oryzae, the causal agent of rice blast, had 89.87% biocontrol efficiency. B. methylotrophicus BC79 colonized rice plant tissues and at 10 days after filtrate application, its population in leaves (1.65 × 108 CFU/g) was much larger than in stems (6.78 × 107 CFU/g) or roots (3.56 × 107 CFU/g). Field trials indicated that BC79 culture filtrate (4000 g/667 m2) showed the highest efficiency for M. oryzae, with 84.8% biocontrol effect, followed by of 15% phenaminomethylacetic acid extract (75.5%) and 20% tricyclazole (76.1%). Seedling and post-transplant stages were the best periods to apply BC79 for control of rice blast. The B. methylotrophicus BC79 strain hence has enormous potential as an agricultural agent for biocontrol of rice blast. © 2012 Elsevier Ltd.

Dai J.,Northwest University, China | Peng H.,Northwest University, China | Chen W.,Northwest University, China | Cheng J.,Shaanxi Tobacco Research Institute | Wu Y.,Northwest University, China
Journal of Applied Microbiology | Year: 2013

Aims: To develop a multiplex real-time PCR assay using TaqMan probes for the simultaneous detection and quantification of Tobacco etch virus (TEV), Potato virus Y (PVY) and Tobacco vein banding mosaic virus (TVBMV). Methods and Results: Specific primer and probe combinations for TEV and TVBMV were developed from the coat protein region of the viral genome. To detect PVY, a primer and probe combination PVY-Univ F, PVY-Univ R and PVY-Univ P for amplifying the coat protein region of the virus genome was employed. The detection limit of multiplex real-time PCR for these viruses was 10 copies μl-1 of the standard plasmid. The multiplex reaction was successful in the detection of these three pathogens, with no non-specific amplification and cross-reaction. Conclusions: This multiplex real-time PCR provides a rapid, effective, specific and sensitive method for the simultaneous detection and quantification of the three pathogens on infected tobacco plants. Significance and Impact of the Study: This multiplex real-time PCR will be useful not only for diagnostic, ecological, epidemiological and pathogenesis studies, but also for investigating host/virus or virus/virus interactions, in particular during mix infection. © 2012 The Society for Applied Microbiology.

Chen W.,Northwest University, China | Liu W.,Northwest University, China | Jiao H.,Northwest University, China | Zhang H.,Northwest University, China | And 2 more authors.
Virologica Sinica | Year: 2014

Tobacco mosaic virus (TMV) causes significant yield loss in susceptible crops irrigated with contaminated water. However, detection of TMV in water is difficult owing to extremely low concentrations of the virus. Here, we developed a simple method for the detection and quantification of TMV in irrigation water. TMV was reliably detected at concentrations as low as 10 viral copies/μL with real-time PCR. The sensitivity of detection was further improved using polyethylene glycol 6000 (PEG6000, MW 6000) to concentrate TMV from water samples. Among the 28 samples from Shaanxi Province examined with our method, 17 were tested positive after virus concentration. Infectivity of TMV in the original water sample as well as after concentration was confirmed using PCR. The limiting concentration of TMV in water to re-infect plants was determined as 102 viral copies/mL. The method developed in this study offers a novel approach to detect TMV in irrigation water, and may provide an effective tool to control crop infection. © 2014 Wuhan Institute of Virology, CAS and Springer-Verlag.

Dai J.,Northwest University, China | Chen W.,Northwest University, China | Cheng J.,Shaanxi Tobacco Research Institute | Wu Y.,Northwest University, China
Journal of Plant Pathology | Year: 2015

An isolate of Tobacco etch virus (TEV-Shaanxi) induces systemic mosaic and necrotic lines or etching in N. Tabacum and is transmitted by M. persicae more efficiently than A. gossypii. The complete genome of TEV-Shaanxi consists of 9,494 nucleotides, excluding the 3’ poly (A) tail. It contains a single large open reading frame (ORF) coding for a polyprotein of 3,054 amino acids, with an AUG start codon and a UGA stop codon. The polyprotein encodes 10 proteins (P1, HC-Pro, P3, 6K1, CI, 6K2, NIa-VPg, NIa-Pro, NIb and CP), whose putative cleavage sites were identified by comparison with sequences of other known potyviruses. The identities of the 10 proteins of TEV-Shaanxi with other members of the genus Potyvirus were 30.5-62.7% and 29.4-68.8% at the nucleotide and amino acid levels, respectively. A short internal open reading frame, PIPO, was also highly conserved. TEV isolates whose sequences are available cluster into two groups which correspond with their host of origin. © 2015, Edizioni ETS. All rights reserved.

Chen W.,Northwest Agriculture and Forestry University | Luo Z.P.,CAS Zhengzhou Research Institute | Liu W.T.,Northwest Agriculture and Forestry University | Liu H.,Northwest Agriculture and Forestry University | And 7 more authors.
Australasian Plant Pathology | Year: 2015

Control of Cucumber mosaic virus (CMV) and Tobacco etch virus is difficult (TEV) mainly because of the lack of resistant tobacco varieties. In this study, the resistance to CMV and TEV of 22 tobacco cultivars, grown under both greenhouse and field conditions, was evaluated from 2007 to 2012. The genetic relationships among these cultivars were also assessed using ISSR methods. Based on the disease index and DAS-ELISA results, one cultivar (Coker86) was characterized as resistant to CMV, and one cultivar (Shuangkang70) was characterized as resistant to TEV. A total of 145 repeatable amplified bands were generated with six ISSR primers, of which 139 were polymorphic. The tobacco cultivars were grouped into five major clusters according to phylogenetic analysis; the similarity coefficient between cultivars ranged from 0.476 to 0.855. This study provides an overall assessment of tobacco cultivars and the results should prove useful for both the control of tobacco virus diseases at present and the improvement of tobacco varieties in future breeding projects. © 2015, Australasian Plant Pathology Society Inc.

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