• Title/Summary/Keyword: Molecular sequence analyses

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Identification and genetic characterization of bacterial isolates causing brown blotch on cultivated mushrooms in Korea

  • Chan-Jung Lee;Hye-Sung Park;Seong-Yeon Jo;Gi-Hong An;Ja-Yun Kim;Kang-Hyo Lee
    • Journal of Mushroom
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    • v.22 no.2
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    • pp.37-47
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    • 2024
  • Fluorescent bacteria were isolated from sporocarps that browned into various mushrooms during survey at places of the production in Korea. We examined the pathogenicity, biodiversity, and genetic characteristics of the 19 strains identified as Pseudomonas tolaasii by sequence analysis of 16S rRNA and White Line Assay. The results emphasize the importance of rpoB gene system, fatty acid profiles, specific and sensitive PCR assays, and lipopeptide detection for the identification of P. tolaasii. As a result of these various analyses, 17 strains (CHM03~CHM19) were identified as P. tolaasii. The phylogenetic analysis based on the 16S rRNA gene showed that all strains were clustered closest to P. tolaasii lineage, two strains (CHM01, CHM02) were not identified as P. tolaasii and have completely different genetic characteristics as a result of fatty acids profile, specific and sensitive PCR, lipopetide detection, rpoB sequence and REP-PCR analysis. Pathogenicity tests showed 17 strains produce severe brown discolouration symptoms to button mushrooms and watersoaking of sporophore tissue within three days after inoculation. But two strains did not produce discolouration symptoms. Therefore, these two strains will be further investigated for correct species identification by different biological and molecular characteristics.

Construction of a full-length cDNA library from Pinus koraiensis and analysis of EST dataset (잣나무(Pinus koraiensis)의 cDNA library 제작 및 EST 분석)

  • Kim, Joon-Ki;Im, Su-Bin;Choi, Sun-Hee;Lee, Jong-Suk;Roh, Mark S.;Lim, Yong-Pyo
    • Korean Journal of Agricultural Science
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    • v.38 no.1
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    • pp.11-16
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    • 2011
  • In this study, we report the generation and analysis of a total of 1,211 expressed sequence tags (ESTs) from Pinus koraiensis. A cDNA library was generated from the young leaf tissue and a total of 1,211 cDNA were partially sequenced. EST and unigene sequence quality were determined by computational filtering, manual review, and BLAST analyses. In all, 857 ESTs were acquired after the removal of the vector sequence and filtering over a minimum length 50 nucleotides. A total of 411 unigene, consisting of 89 contigs and 322 singletons, was identified after assembling. Also, we identified 77 new microsatellite-containing sequences from the unigenes and classified the structure according to their repeat unit. According to homology search with BLASTX against the NCBI database, 63.1% of ESTs were homologous with known function and 22.2% of ESTs were matched with putative or unknown function. The remaining 14.6% of ESTs showed no significant similarity to any protein sequences found in the public database. Gene ontology (GO) classification showed that the most abundant GO terms were transport, nucleotide binding, plastid, in terms biological process, molecular function and cellular component, respectively. The sequence data will be used to characterize potential roles of new genes in Pinus and provided for the useful tools as a genetic resource.

Cloning and Characterization of a Rice cDNA Encoding Glutamate Decarboxylase

  • Oh, Suk-Heung;Choi, Won-Gyu;Lee, In-Tae;Yun, Song-Joong
    • BMB Reports
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    • v.38 no.5
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    • pp.595-601
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    • 2005
  • In this study, we have isolated a rice (Oryza sativa L.) glutamate decarboxylase (RicGAD) clone from a root cDNA library, using a partial Arabidopsis thaliana GAD gene as a probe. The rice root cDNA library was constructed with mRNA, which had been derived from the roots of rice seedlings subjected to phosphorus deprivation. Nucleotide sequence analysis indicated that the RicGAD clone was 1,712 bp long, and harbors a complete open reading frame of 505 amino acids. The 505 amino acid sequence deduced from this RicGAD clone exhibited 67.7% and 61.9% identity with OsGAD1 (AB056060) and OsGAD2 (AB056061) in the database, respectively. The 505 amino acid sequence also exhibited 62.9, 64.1, and 64.2% identity to Arabidopsis GAD (U9937), Nicotiana tabacum GAD (AF020425), and Petunia hybrida GAD (L16797), respectively. The RicGAD was found to possess a highly conserved tryptophan residue, but lacks the lysine cluster at the C-proximal position, as well as other stretches of positively charged residues. The GAD sequence was expressed heterologously using the high copy number plasmid, pVUCH. Our activation analysis revealed that the maximal activation of the RicGAD occurred in the presence of both $Ca^{2+}$ and calmodulin. The GAD-encoded 56~58 kDa protein was identified via Western blot analysis, using an anti-GAD monoclonal antibody. The results of our RT-PCR analyses revealed that RicGAD is expressed predominantly in rice roots obtained from rice seedlings grown under phosphorus deprivation conditions, and in non-germinated brown rice, which is known to have a limited phosphorus bioavailability. These results indicate that RicGAD is a $Ca^{2+}$/calmodulin-dependent enzyme, and that RicGAD is expressed primarily under phosphate deprivation conditions.

Prenatal Diagnosis of Mucolipidosis Type II: Comparison of Biochemical and Molecular Analyses

  • Kosuga, Motomichi;Okada, Michiyo;Migita, Osuke;Tanaka, Toju;Sago, Haruhiko;Okuyama, Torayuki
    • Journal of mucopolysaccharidosis and rare diseases
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    • v.2 no.1
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    • pp.19-22
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    • 2016
  • Purpose: Mucolipidosis type II (ML II), also known as I-cell disease is an autosomal recessive inherited disorder of lysosomal enzyme transport caused by a deficiency of the uridine diphosphate (UDP)-N-acetylglucosamine:lysosomal enzyme N-acetylglucosamine-1-phosphotransferase (GlcNAc-phosphotransferase). Clinical manifestations are skeletal abnormalities, mental retardation, cardiac disease, and respiratory complications. A severely and rapidity progressive clinical course leads to death before 10 years of age. Methods/Results: In this study we diagnosed three cases of prenatal ML II in two different at-risk families. We compared two procedures -biochemical analysis and molecular analysis - for the prenatal diagnosis of ML II. Both methods require an invasive procedure to obtain specimens for the diagnosis. Biochemical analysis requires obtaining cell cultures from amniotic fluid for more than two weeks, and would result in a late diagnosis at 19 to 22 weeks of gestation. Molecular genetic testing by direct sequence analysis is usually possible when mutations are confirmed in the proband. Molecular analysis has an advantage in that it can be performed during the first-trimester. Conclusion: Molecular diagnosis is a preferable method when a prompt decision is necessary.

Characterization of Bacillus luciferensis Strain KJ2C12 from Pepper Root, a Biocontrol Agent of Phytophthora Blight of Pepper

  • Kim, Hye-Sook;Sang, Mee-Kyung;Myung, Inn-Shik;Chun, Se-Chul;Kim, Ki-Deok
    • The Plant Pathology Journal
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    • v.25 no.1
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    • pp.62-69
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    • 2009
  • In this study, we characterized the bacterial strain KJ2C12 in relation with its biocontrol activity against Phytophthora capsici on pepper, and identified this strain using morphological, physiological, biochemical, fatty acid methyl ester, and 16S rRNA gene sequence analyses. Strain KJ2C12 significantly (P=0.05) reduced both final disease severity and areas under the disease progress curves of 5-week-old pepper plants inoculated with P. capsici compared to buffer-treated controls. As for the production of antibiotics, biofilms, biosurfactant, extracellular enzyme, HCN, and swarming activity, strain KJ2C12 produced an extracellular enzyme with protease activity, but no other productions or swarming activity. However, Escherichia coli produced weak biofilm only. Strain KJ2C12 could colonize pepper roots more effectively in a gnotobiotic system using sterile quartz sand compared to E. coli over 4 weeks after treatments. However, no bacterial populations were detected in 10 mM $MgSO_4$ buffer-treated controls. Strain KJ2C12 produced significantly higher microbial activity than the $MgSO_4$-treated control or E. coli over 4 weeks after treatments. Bacterial strain KJ2C12 was identified as Bacillus luciferensis based on morphological, physiological, and biochemical characteristics as well as FAME and 16S rRNA gene sequence analyses. In addition, these results suggested that B. luciferensis strain KJ2C12 could reduce Phytophthora blight of pepper by protecting infection courts through enhanced effective root colonization with protease production and an increase of soil microbial activity.

Neobenedenia girellae infection of aquarium-raised snubnose pompano (Trachinotus blochii) in Korea

  • Nam, U-Hwa;Seo, Hyun-Joon;Hwang, Ilson;Kim, Jeong-Ho
    • Journal of fish pathology
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    • v.33 no.1
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    • pp.15-21
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    • 2020
  • We found skin flukes in snubnose pompano (Trachinotus blochii) from a public aquarium and attempted clear identification of them to the species level by morphology and molecular analyses. Skin flukes were collected from snubnose pompano showing dyspnea, anorexia and mild hemorrhage on the skin. All the fish samples (n=2) were infected with the flukes on the skin, gill and eyes, covered with excessive mucus. The isolated worms were transferred for making slide specimen and PCR amplification targeting 18S rDNA, 28S rDNA, mitochondrial cytochrome c oxidase subunit 1 (mt cox1) and cytochrome b (Cytb) genes for further analyses. Morphology and measurements data of our slide specimen coincided with those of Neobenedenia girellae. The sequence data of 2 genes (28S rDNA and Cytb) and the phylogenetic trees revealed that our specimen consistently belonged to the N. girellae clade. For 18S rDNA and mt cox1 genes, there was no sequence of either of these 2 Neobenedenia species from the type host available in GenBank. This is the first record of N. girellae in snubnose pompano, but it is still unclear if the snubnose pompano is a natural host for N. girellae or not because N. girellae is known to have an unusual broad host range and the host-switching can occur particularly in captive conditions such as aquarium or aquaculture facilities.

Phylogenic Study of Genus Phyllostachys (Phyllostachys) in Korea by Internal Transcribed Spacer Sequence (ITS) (ITS 서열에 의한 한국 왕대속 식물종의 계통분류학적 연구)

  • Lee, Song-Jin;Huh, Man-Kyu;Huh, Hong-Wook;Lee, Byeong-Ryong
    • Journal of Life Science
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    • v.21 no.9
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    • pp.1281-1287
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    • 2011
  • Phyllostachys consists of high and fast growing trees and is a genus in the family Gramineae. The genus has many species in Asia, with main distribution being in India and China. One of the most popular sequences for phylogenetic inference at the generic and infrageneric levels in plants is the internal transcribed spacer (ITS) region of the 18S-5.8S-26S nuclear ribosomal cistron. We evaluated four taxa with the ITS region to estimate phenotypic relationships within the genus Phyllostachys in Korea. Alignment of the DNA sequences required the addition of numerous gaps. Sequence variation within the Phyllostachys was mostly due to natural selection, although several indels and inserts were found. Within the genus Phyllostachys, P. nigra and P. nigra var. henonis were the relatives in the three phylogenetic analyses (MP, ML, and NJ). However, some external nodes were poorly supported. Morphological traits and simple repeats (ISSR) represented the result of a relationship similar to the that of ITS sequences in the genus Phyllostachys. This suggests that ITS sequences are very informative for identification of these taxa.

Development of EST-SSRs and Assessment of Genetic Diversity in Germplasm of the Finger Millet, Eleusine coracana (L.) Gaertn.

  • Wang, Xiaohan;Lee, Myung Chul;Choi, Yu-Mi;Kim, Seong-Hoon;Han, Seahee;Desta, Kebede Taye;Yoon, Hye-myeong;Lee, Yoonjung;Oh, Miae;Yi, Jung Yoon;Shin, Myoung-Jae;Kim, Kyung-Min
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.66 no.4
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    • pp.443-451
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    • 2021
  • Finger millet (Eleusine coracana) is widely cultivated in tropical regions worldwide owing to its high nutritional value. Finger millet is more tolerant against biotic and abiotic stresses such as pests, drought, and salt than other millet crops; therefore, it was proposed as a candidate crop to adapt to climate change in Korea. In 2019, we used expressed sequence tag simple sequence repeat (EST-SSR) markers to evaluate the genetic diversity and structure of 102 finger millet accessions from two geographical regions (Africa and South Asia) to identify appropriate accessions and enhance crop diversity in Korea. In total, 40 primers produced 116 alleles, ranging in size from 135 to 457 bp, with a mean polymorphism information content (PIC) of 0.18225. Polymorphism was detected among the 40 primers, and 13 primers were found to have PIC values > 0.3. Principal coordinate and phylogenetic analyses, based on the combined data of both markers, grouped the finger millet accessions according to their respective collection areas.Therefore, the 102 accessions were classified into two groups, one from Asia and the other from Africa. We have conducted an in-depth study on the finger millet landrace pedigree. By sorting out and using the molecular characteristics of each pedigree, it will be useful for the management and accession identification of the plant resource. The novel SSR markers developed in this study will aid in future genetic analyses of E. coracana.

Phylogenomics and its Growing Impact on Algal Phylogeny and Evolution

  • Adrian , Reyes-Prieto;Yoon, Hwan-Su;Bhattacharya, Debashish
    • ALGAE
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    • v.21 no.1
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    • pp.1-10
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    • 2006
  • Genomic data is accumulating in public database at an unprecedented rate. Although presently dominated by the sequences of metazoan, plant, parasitic, and picoeukaryotic taxa, both expressed sequence tag (EST) and complete genomes of free-living algae are also slowly appearing. This wealth of information offers the opportunity to clarify many long-standing issues in algal and plant evolution such as the contribution of the plastid endosymbiont to nuclear genome evolution using the tools of comparative genomics and multi-gene phylogenetics. A particularly powerful approach for the automated analysis of genome data from multiple taxa is termed phylogenomics. Phylogenomics is the convergence of genomics science (the study of the function and structure of genes and genomes) and molecular phylogenetics (the study of the hierarchical evolutionary relationships among organisms, their genes and genomes). The use of phylogenetics to drive comparative genome analyses has facilitated the reconstruction of the evolutionary history of genes, gene families, and organisms. Here we survey the available genome data, introduce phylogenomic pipelines, and review some initial results of phylogenomic analyses of algal genome data.

New Record of Scolecenchelys fuscogularis (Anguilliformes: Ophichthidae) Leptocephali from Korea, as Revealed by Morphological and Molecular Analyses

  • Ji, Hwan-Sung;Kim, Jin-Koo;Lee, Soo Jeong;Kimura, Seishi;Hibino, Yusuke
    • Fisheries and Aquatic Sciences
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    • v.18 no.1
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    • pp.109-113
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    • 2015
  • Three leptocephali (22.2, 22.7, 56.0 mm in total length) collected from the East/Japan Sea were identified by morphological and genetic analyses as belonging to the genus Scolecenchelys (Anguilliformes, Ophichthidae). Morphologically, the specimens were characterized by 148-158 myomeres, 10 gut swellings, dorsal fin origin above middle of the body, and 6 postanal melanophores between the anus and the caudal margin. An analysis of an 849-base pair 12S rRNA sequence of mitochondrial DNA showed that sequences are concordant with those of adult Scolecenchelys fuscogularis (genetic distance = 0.001). Furthermore total number of myomeres is consistent with the total number of vertebrae in adult S. fuscogularis. This study provides the first description of the morphological characteristics of S. fuscogularis leptocephali and their variations with size. The Korean name of S. fuscogularis is "Ga-neun-mul-baem", established by Ji et al. (2012).