• The Plant Pathology Journal
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• 제18권2호
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• pp.98-101
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• 2002

Using phytoplasma universal primer pair Pl and P7, a fragment of about 1.8 kb nucleotide sequences of 16S rRNA gene and 16S-23S rRNA intergenic spacer region, and a portion of 23S rRNA gene of jujube witches'broom (JWB) and mulberry dwarf(MD) phytoplasmas were determined. The nucleotide sequences of JWB and MD were 1,850 bp and 1,831 bp long, respectively. The JWB phytoplasma sequence was aligned with the homologous sequence of MD phytoplasma. Twenty-eight base insertions and nine base deletions were found in the JWB phytoplasma sequence compared with that of MD phytoplasma. The similarity of the aligned sequences of JWB and MD was 84.8%. The near-complete 16S rRNA gene DNA sequences of JWB and MD were 1,529 bp and 1,530 bp in length, respectively, and revealed 89.0% homology. The 16S-23S rRNA intergenic spacer region DNA sequences were 263 bp and 243 bp in lengths respectively, while homology was only 70% and the conserved tRNA-lle gene of JWB and MD was located into the intergenic space region between 16S-23S rRNA gene. The nucleotide sequences were 77 bp long in both JWB and MD, and showed 97.4% sequence homology. Based on the phylogenetic analysis of the two phytoplasmas, the JWB phytoplasma belongs to the Elm yellow phytoplasma group (16S rV), whereas, the MD phytoplasma belongs to the Aster yellow group (16S rI).

• The Plant Pathology Journal
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• 제28권2호
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• pp.191-196
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• 2012

Periwinkle seedlings (Cantharanthus roseus) were inoculated with jujube witches'- broom (JWB) phytoplasma via grafting to analyze the migration of JWB phytoplasmas within the host plant. The phytoplasmas were detected using nested polymerase chain reaction (PCR) and fluorescence microscopy. Fluorescence microscopy was a simple and easy method of detecting phytoplasmas; however, it was not sufficiently sensitive to detect very low phytoplasma concentrations. Therefore, the migration of JWB phytoplasma was investigated through PCR. The first migration of JWB phytoplasma from an infected tissue to healthy tissues occurred late. After grafting, the phytoplasmas moved from the inoculated twig (or scion) to the main stem, which took 28 days. Afterward, the phytoplasma migrated faster and took less than 4 days to spread into the roots from the main stem. All twigs were then successively colonized by the JWB phytoplasmas from the bottom to the top. JWB phytoplasma was detected via nested PCR in all parts of the periwinkle seedling 82 days after inoculation. Based on these results, the inoculated JWB phytoplasma appeared to migrate downward to the roots along the main stem during the early stages, and then continued to move upward, colonizing twigs along the way until they reached the apex.

• 한국식물병리학회:학술대회논문집
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• 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
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• pp.136.2-137
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• 2003

In order to diagnose and differentiate jujube witches' broom (JWB) phytoplasma rapidly, oligonucleotide primer pair, 16Sr(V) F/R, for polymerase chain reactions (PCRs) was designed on the basis of 165 rRNA sequences of JWB phytoplasma. The PCR employing phytoplasma universal primer pair P1/P7 consistently amplified DNA in all tested phytoplasma isolates. But no phytoplasma DNA was detected in healthy jujube seedlings. The nested PCR, the primer pair 16S(V) F/R, about 460 bp fragment, amplified DNA in all tested JWB and related phytoplasmas including LiWB phytoplasma of the 165 rRNA group V, but no DNA amplification was detected from other phytoplasma strains such as group 16SrI (Aster yellows) and group 16SrⅩII (Stolbur group) phytoplasmas in which mulberry dwarf phytoplasma and chrysanthemum witches broom phytoplasma are belonged to, respectively The same results were obtained from both Korean- and Chinese-isolates of JWB. Nested-PCR using phytoplasma universal primer pair P1/P7 and 16S rRNA group V specific primer pair 16S(V) F/R could detect group V phytoplasma rapidly and easily, in particular JWB phytoplasma.

• The Plant Pathology Journal
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• 제21권1호
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• pp.55-58
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• 2005

In order to diagnose and differentiate jujube witches' broom (JWB) phytoplasma rapidly, oligonucleotide primer pair, 16Sr(V) F/R, for polymerase chain reactions (PCRs) was designed on the basis of 16S rRNA sequences of JWB phytoplasma. The PCR employing phytoplasma universal primer pair P1/P7 consistently amplified DNA in all tested phytoplasma isolates. But no phytoplasma DNA was detected from healthy jujube seedlings. The nested PCR, the primer pair 16S(V) F/R, about 460 bp fragment, amplified DNA in all tested JWB and related phytoplasmas including ligustrum witches' broom phytoplasma of the 16S rRNA group V, but no DNA amplification was detected from other phytoplasma strains such as groups 16SrI (Aster yellows) and 16SrXII (Stolbur group) in which mulberry dwarf phytoplasma and chrysanthemum witches' broom phytoplasma belong to, respectively. The same results were obtained from both Korean and Chinese isolates of JWB phytoplasma. Nested-PCR using phytoplasma universal primer pair P1/P7 and 16SrV group-specific primer pair 16S(V) F/R could detect group V phytoplasmas rapidly and easily, in particular JWB phytoplasma.

• 한국산림과학회지
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• 제95권6호
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• pp.631-635
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• 2006

파이토플라스마 증폭 primer, R16F2n/R2를 이용하여 뽕나무 오갈병 파이토플라스마와 대추나무 빗자루병 파이토플라스마에 대하여 SSCP분석법을 이용하여 염기변이 분석을 하였다. 그 결과 뽕나무 및 대추나무 파이토플라스마는 약 1.2 kb PCR 산물을 이용하더라도 뚜렷한 밴드차이를 나타내었다. 유사한 SSCP밴드 패턴을 보이는 두 시료 간의 밴드형태를 뚜렷하게 구별하는 방법을 찾기 위하여 뽕나무 오갈병 파이토플라스마와 대추나무 빗자루병 파이토플라스마의 PCR산물을 혼합한 후 SSCP분석 결과, 전기영동상에서 대추나무 파이토플라스마와 뽕나무 파이토플라스마의 SSCP 밴드패턴 모두를 관찰할 수 있었다. 본 연구 결과, 기존에 약 600bp 크기로 한정된 것으로 알려진 SSCP 분석을 PCR 산물 1.2 kb을 이용하여 유사한 SSCP 밴드패턴을 보이는 두 시료간의 밴드형태를 두 시료의 PCR 산물을 혼합하여 SSCP분석함으로써 뚜렷하게 구별할 수 있었다.

• The Plant Pathology Journal
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• 제18권6호
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• pp.308-312
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• 2002

Heteroduplex mobility assay (HMA) and single-strand conformation polymorphism (SSCP) analyses combined with PCR were developed for genetic differentiation of various phytoplasma isolates. In the HMA and SSCP analyses, differences in the mobility shifts and the SSCP band patterns identified three distinct types of phyto-plasmas: Type Ⅰ, jujube witches'-broom (JWB) and ligustrum witches'-broom (LiWB); Type Ⅱ, mulberry dwarf（MD) and sumac witches'-broom (SuWB); and Type Ⅲ, paulownia witches'-broom (PaWB). Results of the sequence analyses revealed that phytoplasmas of JWB and MD had 100％ homology with LiWB and SuWB, respectively. On the other hand, PaWB phyto-plasma had 97.8% homology with MD phytoplasma. The PCR-HMA and SSCP techniques were very useful in determining variations in sequence among several isolates of phytoplasmas. Furthermore, the methods were rapid, economical, highly sensitive, and easy to handle with the gels.

• 식물병연구
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• 제26권1호
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• pp.19-28
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• 2020

대추나무 빗자루병은 1950년대에 남한지역에 대발생하여 대추 재배지를 황폐화시킨 고위험 식물병이다. 최근 전국적으로 대추 재배면적이 크게 증가하고 있어 빗자루병 대발생 위험성을 사전에 파악하기 위한 기초자료를 얻고자 2017년과 2019년에 걸쳐 보은 지역의 대추 과원에서 병조사를 하였다. 또한, 빗자루병 조기 진단에 활용할 수 있는 다양한 병징을 찾아내고, 성보싸이클린수화제(OTC, 옥시테트라사이클린칼슘알킬트리메틸암모늄 수화제, 유효성분 17%)의 수간주사 방제효과 증진을 위한 연구를 수행하였다. 그 결과 현재까지 충북 보은 지역의 대추나무 빗자루병 발병률(이병주율)은 매우 낮았으나, 발생 지역과 발생 농가를 중심으로 점차 의미 있게 증가하고 있었다. OTC 방제시험 결과 처리 시기는 대추나무 생육 초기가 가장 효과적이었다. 그러나 발병이 심한 나무에서는 수간주사 후 당해 연도에도 같은 나무의 다른 건전한 가지에서 병징이 발현되었다. 6월에 1차 수간주사를 한 처리에서 2차 수간주사는 5개월 후보다 한 달 후에 하였을 때 방제효과가 더 좋았다. 이러한 결과는 조기 병징을 활용한 진단과 이를 근거로 한 조기 방제가 빗자루병 피해를 사전에 차단하고 병 방제효과를 높이는 데 매우 효과적일 수 있음을 시사한다.

• 한국산림과학회지
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• 제102권2호
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• pp.223-228
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• 2013

파이토플라스마 증폭 프라이머, P1/P7 및 R16F2n/R2를 이용하여 뽕나무 품종 42개체에 대하여 뽕나무오갈병 파이토플라스마의 전염여부를 조사한 결과 공시 모두에서 파이토플라스마가 검출되었다. 뽕나무오갈병 파이토플라스마 단일염기변이를 SSCP분석기법을 응용하여 분석조건을 조사한 결과 P1/P7(약 1.8 kb) 및 R16F2n/R2(약 1.2kb)로 증폭한 PCR산물에서는 6% polyacrylamide gel 농도, 150V, $10^{\circ}C$의 전기영동 조건에서 SSCP밴드패턴이 나타났다. 유사한 SSCP밴드 패턴을 보이는 두 시료간의 밴드형태를 뚜렷하게 구별하는 방법을 찾기 위하여 뽕나무 오갈병파이토플라스마와 대추나무 빗자루병 파이토플라스마의 P1/P7 및 R16F2n/R2 프라이머로 증폭한 PCR산물을 혼합한 후 SSCP분석 결과, 전기영동상에서 대추나무 파이토플라스마와 뽕나무 파이토플라스마의 SSCP 밴드패턴 모두를 관찰할 수 있었다. 본 연구 결과, 기존에 약 600 bp 크기로 한정된 것으로 알려진 SSCP 분석법을 응용하여 파이토플라스마 PCR 산물 1.8 kb 또는 1.2 kb 크기에서도 유사한 SSCP 밴드패턴에 의하여 단일염기변이를 검출할 수 있었다.