• Korean Journal Plant Pathology
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• v.14 no.6
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• pp.636-641
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• 1998

Digoxigenin (DIG) was used to prepare nucleic acid probe for the detection of RNA of potato leafroll virus (PLRV) in the potato leaf extracts. The 0.6 kb coat protein (CP) gene cDNA of PLRV in plasmid pSPT 18 vector was labeled with digoxigenin by in vitro run-off transcription and then used for cRNA probe. In the several buffers tested for increase the total RNA extraction efficiency AMES buffer was the most suitable for this detection method. The RNA extracts from potato leaves shown symptoms of PLRV were dot blotted onto nylon membrane and hybridized with labeled RNA probes. After hybridization, labeled RNA bound to PLRV RNA on membrane was detected with anti-digoxigenin alkaline phosphatase. 5-bromo-4-chloro-3-indolyl-phosphate/nitroblue tetrazolium (NBT) salt and CSPD were used as substrate for colorimetric and film exposure detection, respectively. These detection methods were very sensitive allowing for detection of 1/32 diluted total RNA extract from 100 mg leaf tissue.

• Journal of Plant Biotechnology
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• v.32 no.4
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• pp.243-250
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• 2005

The coat protein gene (AF296280) of the Korean isolate Potato leafroll virus (PLRV) was cloned and the open reading frame (627 bp) was transformed into potato (Solanum tuberosum cv. Superior). Out of seventeen individual transgenic lines, five lines were identified to confer resistance to PLRV through the five generation's selection program in the greenhouse as well as isolated trial field. Successful introduction and genetic stability of coat protein gene in the genome of potato were confirmed by polymerase chain reaction (PCR), Southern blot hybridization and northern blot hybridization. Some of the transgenic lines were highly resistant to PLRV but did not show any resistance to less homologous Potato virus Y (PVY). Our results suggest that the resistance to PLRV is due to homology dependent gene silencing by sense strand coat protein gene. In addition, the results of field test through five generations showed that there were no significant differences comparing to nontransgenic potatoes in the morphological aspect of shoot as well as tuber, Ho remarkable differences were also observed in the major agronomic characters and yields except for the resistance to PLRV.

• The Plant Pathology Journal
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• v.33 no.5
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• pp.522-527
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• 2017

We determined the effects of atmospheric temperature ($10-30{\pm}2^{\circ}C$ in $5^{\circ}C$ increments) and carbon dioxide ($CO_2$) levels ($400{\pm}50ppm$, $540{\pm}50ppm$, and $940{\pm}50ppm$) on the infection of Solanum tuberosum cv. Chubaek by Potato leafroll virus (PLRV). Below $CO_2$ levels of $400{\pm}50ppm$, the PLRV infection rate and RNA content in plant tissues increased as the temperature increased to $20{\pm}2^{\circ}C$, but declined at higher temperatures. At high $CO_2$ levels ($940{\pm}50ppm$), more plants were infected by PLRV at $30{\pm}2^{\circ}C$ than at 20 or $25{\pm}2^{\circ}C$, whereas PLRV RNA content was unchanged in the $20-30{\pm}2^{\circ}C$ temperature range. The effects of atmospheric $CO_2$ concentration on the acquisition of PLRV by Myzus persicae and accumulation of PLRV RNA in plant tissues were investigated using a growth chamber at $20{\pm}2^{\circ}C$. The M. persicae PLRV RNA content slightly increased at elevated $CO_2$ levels ($940{\pm}50ppm$), but this increase was not statistically significant. Transmission rates of PLRV by Physalis floridana increased as $CO_2$ concentration increased. More PLRV RNA accumulated in potato plants maintained at 540 or $940{\pm}50ppm$ $CO_2$, than in plants maintained at $400{\pm}50ppm$. This is the first evidence of greater PLRV RNA accumulation and larger numbers of S. tuberosum plants infected by PLRV under conditions of combined high $CO_2$ levels ($940{\pm}50ppm$) and high temperature ($30{\pm}2^{\circ}C$).

• Korean journal of applied entomology
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• v.41 no.4
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• pp.247-253
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• 2002

Based on accumulated data during 1977-2001, seasonal fluctuations of migrated aphids at Daegwallyeong highlands were analyzed. In addition, rates of PLRV transmission by migrated aphids were investigated by inoculation on indicator plant, Physazis floridana, and ELISA in 2000-2001, and the change of PLRV transmission rates by aphids was compared with that of 1989-1991. The average migrated aphid population densities in 1976-1980, 1991-1995, and 1996-2000 were 575.2, 2959.4 and 2281.6, respectively, showing gradual increase in recent years. The average peak time of aphid migration was from early to mid June during 1977-2001, showing any significant differences over the years. The dominant species, however, changed slowly; before mid 1980s M. persicae flied dominantly, but after mid 1980s Aphis gossypii did. Hahm et al. (1991) reported that PLRV transmission rate of migrated aphids during 1989-1991 was 6.7-10.0%. In 2000-2001, however, migrated aphids at Daegwallyeong highland showed 10.1-11.0%. Although present PLRV transmission rate was slightly higher than that of 10 years ago, taking increased population densities and diversity of migrated aphids into account, there was no significant change of PLRV transmission rate over the years.

• The Plant Pathology Journal
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• v.18 no.1
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• pp.1-5
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• 2002

The availability of nucleotide sequences of the coat protein gene of Potato leafroll virus (PLRV) permitted the construction of DNA primers that were utilized for cDNA synthesis. Polymerase chain reaction (PCR) products of a 487 bp. and approximately 500 bp DNA fragments were amplified from nucleic acid extracts of PLRV-infected tissue and strawberry mild yellow-edge (SMYE) diseased strawberry tissue, respectively. The amplified DNA fragments were further differentiated by hybridization analysis with a CDNA probe for the coat protein gene of PLRV and restriction fragment length polymorphism (RFLP) analysis. These results suggest that a luteovirus is associated with the SMYE disease.

• The Plant Pathology Journal
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• v.27 no.4
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• pp.385-389
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• 2011

A new reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for the Potato leafroll virus (PLRV) was developed and compared with conventional reverse transcription polymerase chain reaction (RT-PCR) to address its advantages over RTPCR. RT-LAMP primers were designed from the open reading frame 3 (ORF3) sequence of PLRV. The RT-LAMP reactions were conducted without or with a set of loop primers. By real-time monitoring using Turbimeter, the RT-LAMP (with loop primers) detects PLRV in less than 30 min, compared to 120 min of RT-PCR. By adding fluorescent reagent during the reaction, final products of the RT-LAMP were fluorescently visualized under UV light or could be differentiated by naked-eye inspection under normal light. The RT-LAMP was extremely sensitive, about 2000-fold more sensitive than RT-PCR. This study presents great potential of the RT-LAMP for diagnosis and PLRV epidemiology because RT-LAMP method is speedy, sensitive, inexpensive, and convenient.

• Korean Journal Plant Pathology
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• v.11 no.2
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• pp.101-106
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• 1995

Deep blue fluorescence of resorcin blue-stained callose was observed only in the potato leafroll virus (PLRV)-infected potato plants, but not in other potato viruses investigated. The plant sections stained with aniline blue showed non-specific fluorescence regardless of PLRV infection, which means that aniline blue is not suitable for the staining of callose induces by PLRV infection. The fluorescence of resorcin blue-stained callose was more easily detectable than autofluorescence by a direct fluorescence detection method because of its deep blue color. The lateral branch of lower leaves was turned out to be the best material for fluorescence observation of all plant parts tested. In comparison of diagnostic efficacy of this technique to enzyme-linked immunosorbent assay (ELISA), PLRV infected potato plants showed corresponding increment of the fluorescence of resorcin blue stained callose as absorption values in ELISA increased. In the future, the criteria measuring the fluorescence objectively are thought to be determined for the practical application to the diagnosis of potato leafroll disease.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.59 no.4
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• pp.498-504
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• 2014

Potato virus Y (PVY) and potato leafroll virus (PLRV) are among the most damaging potato viruses and prevalent in most potato growing areas. In this study, cryopreservation was used to eradicate PVY and PLRV using two cryogenic methods. Potato shoot tips proliferated in vitro were cryopreserved through droplet-vitrification and encapsulation-vitrification using plant vitrification solution 2 (PVS2; 30% glycerol + 15% dimethyl sulfoxide + 15.0% ethylene glycol + 13.7% sucrose) and modified PVS2. Both cryogenic procedures produced similar rates of survival and regrowth, which were lower than those from shoot tip culture alone. The health status of plantlets regenerated from shoot tip culture alone and cryopreservation was checked by reverse transcription-polymerase chain reaction. The frequency of virus-free plants regenerated directly from highly proliferating shoot tips reached 42.3% and 48.6% for PVY and PLRV, respectively. In comparison, the frequency of PVY and PLRV eradication after cryopreservation was 91.3~99.7% following shoot-tip culture. The highest cryopreserved shoot tip regeneration rate was observed when shoot tips were 1.0~1.5 mm in length, but virus eradication rates were very similar (96.4~99.7%), regardless of shoot tip size. This efficient cryotherapy protocol developed to eliminate viruses can also be used to prepare potato material for safe long-term preservation and the production of virus-free plants.

• The Plant Pathology Journal
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• v.32 no.4
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• pp.321-328
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• 2016

We examined the effects of temperature on acquisition of Potato virus Y-O (PVY-O), Potato virus A (PVA), and Potato leafroll virus (PLRV) by Myzus persicae by performing transmission tests with aphids that acquired each virus at different temperatures. Infection by PVY-O/PVA and PLRV increased with increasing plant temperature in Nicotiana benthamiana and Physalis floridana, respectively, after being transmitted by aphids that acquired them within a temperature range of $10-20^{\circ}C$. However, infection rates subsequently decreased. Direct qRT-PCR of RNA extracted from a single aphid showed that PLRV infection increased in the $10-20^{\circ}C$ range, but this trend also declined shortly thereafter. We examined the effect of temperature on establishment of virus infection. The greatest number of plants became infected when N. benthamiana was held at $20^{\circ}C$ after inoculation with PVY-O or PVA. The largest number of P. floridana plants became infected with PLRV when the plants were maintained at $25^{\circ}C$. PLRV levels were highest in P. floridana kept at $20-25^{\circ}C$. These results indicate that the optimum temperatures for proliferation of PVY-O/PVA and PLRV differed. Western blot analysis showed that accumulations of PVY-O and PVA coat proteins (CPs) were lower at $10^{\circ}C$ or $15^{\circ}C$ than at $20^{\circ}C$ during early infection. However, accumulation increased over time. At $25^{\circ}C$ or $30^{\circ}C$, the CPs of both viruses accumulated during early infection but disappeared as time passed. Our results suggest that symptom attenuation and reduction of PVY-O and PVA CP accumulation at higher temperatures appear to be attributable to increased RNA silencing.

• Korean journal of applied entomology
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• v.57 no.4
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• pp.415-423
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• 2018

Aphid is a typical vector that transfers various kinds of viruses to potatoes. Therefore, it is very important to control aphids moving into potato fields. We investigated the seasonal movement pattern of aphids and its virus transmission rates mainly in the three seed potato production regions at highland in Gangwon-do, Korea. In addition, we identified the aphid species with over-wintering eggs collected from barks or twigs of total 57 tree species around potato fields in winter season. The peak time of summer and winter migration of aphid was at the mid-June and the early October, respectively. A 2.8% of total aphid trapped in yellow water-pan trap was turned out PLRV-borne, and the virus transmission rate was 15.4% by Myzus persicae and 9.1% by Macrosiphum euphorbiae. PLRV-borne aphids started to flow in from the late May, and virus transmission rate of aphid trapped in mid-June was the highest with 10.4%. Totally 14 species of aphid eggs wintered in the 17 species of trees including Acer pictum subsp. mono and Acer pseudosieboldianum at the 11 sites. In particular, because it is not certain that Betula platyphylla var. japonica and Yamatocallis hirayamae do transmit potato virus, but they over-wintered in host plants distributed over a wide area, further research on transmission ability is necessary.