• Title/Summary/Keyword: internal transcribed spacer-restriction fragment length polymorphism (ITS-RFLP)

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Use of Restriction Fragment Length Polymorphism Analysis to Differentiate Fungal Strains in Sunchang Meju

  • Jung, Jong-Hyun;Seo, Dong-Ho;Bhoo, Sung-Hee;Ha, Suk-Jin;Kim, Jong-Sang;Kim, Jeong-Hwan;Kwon, Dae-Young;Cha, Jae-Ho;Park, Cheon-Seok
    • Food Science and Biotechnology
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    • v.17 no.4
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    • pp.888-891
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    • 2008
  • Twenty-three fungal strains were isolated from meju that had originated from the Sunchang province, the famous location for making fermented soybean foods in Korea. The restriction fragment length polymorphism (RFLP) of the internal transcribed spacer (ITS) region of the rDNA (ITS-RFLP) was applied to differentiate the isolated fungal strains. First, the ITS region by polymerase chain reaction (PCR) with specific primers was amplified and then cleaved the products with different restriction enzymes. Cleavage of the amplified fragments with the restriction enzymes AluI, HaeIII, HhaI, and TaqI revealed extensive polymorphisms. The ITS-RFLP results highly correlated with ITS sequence analysis. All of the 23 fungal strains were classified into 5 groups by ITS-RFLP analysis. Aspergillus oryzae was the major fungal strain isolated from Sunchang meju (12 out of 23), while Aspergillus fumigatus was the next most frequently isolated strain (7 out of 23). In contrast, it was found that Fusarium asiaticum, Aspergillus sydowii, and Arthrinium sp. were the minor fungal strains in meju.

PCR-RFLP and Sequence Analysis of the rDNA ITS Region in the Fusarium spp.

  • Min, Byung-Re;Lee, Young-Mi;Choi, Yong-Keel
    • Journal of Microbiology
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    • v.38 no.2
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    • pp.66-73
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    • 2000
  • To investigate the genetic relationship among 12 species belonging to the Fusarium section Martiella, Dlaminia, Gibbosum, Arthrosporiella, Liseola and Elegans, the internal transcribed spacer(ITS) regions of ribosomal DNA (rDNA) were amplified with primer pITS1 and pITS4 using the polymerase chain reaction(PCR). After the amplified products were digested with 7 restriction enzymes, restriction fragment length polymorphism (RFLP) patterns were analyzed. The partial nucleotide sequences of the ITS region were determined and compared. Little variation was observed in the size of the amplified product having sizes of 550bp or 570bp. Based on the RFLP analysis, the 12 species studied were divided into 5 RFLP types. In particular, strains belonging to the section Martiella were separated into three RFLP types. Interestingly, the RFLP type of F. solani f. sp. piperis was identical with that of isolates belonging to the section Elegans. In the dendrogram derived from RFLP analysis of the ITS region, the Fusarium spp. examined were divided into two major groups. In general, section Martiella excluding F. solani f. sp. piperis showed relatively low similarity with the other section. The dendrogram based on the sequencing analysis of the ITS2 region also gave the same results as that of the RFLP analysis. As expected, 5.8S, a coding region, was highly conserved, whereas the ITS2 region was more variable and informative. The difference in the ITS2 region between the length of F. solani and its formae speciales excluding F. solani f. sp. piperis and that of other species was caused by the insertion/deletion of nucleotides in positions 143-148 and 179-192.

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Genetic characterization of Phellinus baumii PMO-P4 by analyzing restriction fragment length polymorphisms of nuclear ribosomal DNA internal transcribed spacers (ITS) (Ribosomal DNA의 ITS부위에 대한 RFLP 분석에 의한 Phellinus baumii PMO-P4의 유전학적 특성)

  • Chang, Yun-Hee;Kim, Tae-Rack;Kim, Hyun-Su;Yeo, Ik-Hyun;Lee, Sang-Youn;Ha, Hyo-Cheol
    • Journal of Mushroom
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    • v.4 no.2
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    • pp.43-47
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    • 2006
  • PMO-P4, being cultivated as "Sanghwang" in Korea, was proved to be P. baumii based on ITS (internal transcribed spacer) sequencing and RFLP (Restriction Fragment Length Polymorphism) patterns along with some Phellinus species including P. linteus. The similaraty of ITS sequencing between PMO-P4 and other Phellinus species was given the range of 48.6%~72.2%, showing the highest homology from P. linteus and the lowest from P. gilvus.

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Identification of Korean Suminoe Oyster (Crassostrea ariakensis) by RFLP Analysis

  • Kim, Mi-Jung;Park, Jung-Youn;Allen, Stanish K.;An, Hye-Suck
    • Fisheries and Aquatic Sciences
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    • v.11 no.1
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    • pp.32-35
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    • 2008
  • The Suminoe oyster, Crassostrea ariakensis, occurs in estuaries where rivers meet seawater. In Korea, it is one of the most popular fisheries resources in the Nam River and Sumjin River. However, the genetic identification of this species has been questioned, because specimens are often mis-identified as other species. To identify the species, we conducted polymerase chain reaction (PCR) amplification of the internal transcribed spacer-1 (ITS-1) region, followed by digestion with the restriction enzyme HaeIII. Restriction profiles for oysters collected from Korea, Japan, and China (north and south) were determined by comparing the PCR-restriction fragment length polymorphism (RFLP) patterns of the ITS-1 regions. Our study verified that the oysters collected from Korea were C. ariakensis based on the PCR-RFLP patterns. These results emphasize the value of molecular markers for identifying morphologically uncertain species.

The Relative Identification of C. officinale and L. chuanxiong by PCR-Mediated Fingerprinting (천궁류(川芎類) 한약재의 유전자 감식 연구)

  • Choi, Ho-Young;Kim, Dong-Wook;Kim, Dong-Eun;Suh, Young-Bae;Ham, In-Hye
    • The Korea Journal of Herbology
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    • v.20 no.4
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    • pp.151-161
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    • 2005
  • Objectives : Our research purpose is to establish the standard identification analysis on C. officinale and L. chuanxiong in Korea and China by PCR-mediated fingerprinting. Methods : The Restriction Fragment Length Polymorphism (RFLP) and Randomly Amplified Polymorphic DNA (RAPD) method was used on Internal Transcribed Spacer (ITS) regions and rbcL regions to compare and discriminate genes extracted from crude drugs as C. officinale and L. chuanxiong in Korea and China. Results : L. chuanxiong Korea and China have very similar polymorphism, whereas L. chuanxiong in Korea and C. officinale have very different polymorphism in RFLP. And restriction enzymes AluI and SacI forms the specific fragment band only in C. officinale, they can be used as RFLP marker on ITS regions to discriminate among the species. Conclusions : The results could be applied in discriminating crude drugs among C. officinale and L. chuanxiong in Korea and China. Also they could be used in controlling drug quality, preserving medicinal plants, and improving plant description.

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Analysis of Molecular Diversity in Castanopsis sieboldii with Felt Disease Caused by Septobasidium sp. (Septobasidium sp.에 의한 구실잣밤나무 고약병의 분자학적 다양성 분석)

  • Geon-Woo Lee;Sang-Tae Seo;Byeongjin Cha;Sang-Sub Han
    • Research in Plant Disease
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    • v.29 no.4
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    • pp.420-424
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    • 2023
  • In 2020, within the Dongbaekdongsan area in Jeju Island, a Septobasidium sp. associated with a felt disease in Castanopsis sieboldii (Makino) Hatus. ex T. Yamaz. & Mashiba was identified. The symptom included the presence of brown, thin, and silk-like mycelial mats attached to the tree's bark, displaying variations in size from large to small. To induce hyphal growth, the samples collected were incubated in a moist chamber, and the newly formed hyphae were subjected to genomic DNA extractions. The nucleotide sequences of the internal transcribed spacer and small subunit rDNA genes were determined, and molecular characteristics among the isolates were investigated through polymerase chain reaction-based restriction fragment length polymorphism analysis. This Septobasidium sp. exhibited distinct morphological and phylogenetic features compared to those that were previously reported in South Korea. Consequently, this strain is taxonomically classified as a provisionally novel species of Septobasidium. Furthermore, the observed felt disease exhibited a high degree of host specificity, as it was exclusively identified in C. sieboldii without occurrence in other tree species at the time of observation.

RAPD Analysis on the Species of Pinelliae Tuber (RAPD 방법을 이용한 반하류 한약재의 감별 연구)

  • 배명효;김규열;정유헌;최호영
    • The Journal of Korean Medicine
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    • v.20 no.4
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    • pp.16-22
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    • 2000
  • This study intends to report the significance of several experimental results obtained from analysing the genes extracted from the plants and herbal medicine such as P. temalta (Thunb.) Breit, A. amurense var serratum Nakai, A. erubescens (Wall.) Schott, Pinelliae Tuber and Arisaematis Tuber, mainly by the method of RAPD(randomly amplified polymorphic DNA) and the method of RFLP(restriction fragment length polymorphism) on ITS(internal transcribed spacer) region. Genomic DNA could be extracted from both original plants and dried materials. DNA fragments of P. temata kind and A. amurense kind showed the same aspect separately within the same species under the method of RAPD using random primer, while various aspects(polymorphism) were discovered among different species. In RAPD analysis by uniprimer, common bands were extracted from all types of P. temata in the case of uniprimer #4, which were distinguished from the kind of A. amurense. Other polymorphic bands appeared in between different A. amurense species as well. In the case of uniprimer #11, particular band came out in the kind of P. temata. On the other hand, in the case of uniprimer #5, #6, and #8, various bands(polymorhism) were revealed in both kinds of P. temata and A. amurense. Although further study is needed to ascertain whether these results are due to the differences of species, kinds, or growing place, the results could be used as a scientific method of identifying the substitutes for A. amurense genus. The author believes that as if P. temata class of plants used in this experiment are different among themselves in terms of the shape, size and property, those are clearly a class of P. temata or belong to the same genus.

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Development of Species-specific Primers for Rapid Detection of Phellinus linteus and P. baumii

  • Kim, Mun-Ok;Kim, Gi-Young;Nam, Byung-Hyouk;Jin, Cheng-Yun;Lee, Ki-Won;Park, Jae-Min;Lee, Sang-Joon;Lee, Jae-Dong
    • Mycobiology
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    • v.33 no.2
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    • pp.104-108
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    • 2005
  • Genus Phellinus taxonomically belongs to Aphyllophorales and some species of this genus have been used as a medicinal ingredients and Indian folk medicines. Especially, P. linteus and morphological-related species are well-known medicinal fungi that have various biological activities such as humoral and cell-mediated, anti-mutagenic, and anti-cancer activities. However, little is known about the rapid detection for complex Phellinus species. Therefore, this study was carried out to develop specific primers for the rapid detection of P. linteus and other related species. Designing the species-specific primers was done based on internal transcribed spacer sequence data. Each primer set detected specifically P. linteus (PL2/PL5R) and P. baumii (PB1/PB4R). These primer sets could be useful for the rapid detection of specific-species among unidentified Phellinus species. Moreover, restriction fragment length polymorphism analysis of the ITS region with HaeIII was also useful for clarifying the relationship between each 5 Phellinus species.