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EST Profiling for Seed-hair Characteristic and Development of EST-SSR and SNP Markers in Carrot (당근 종모 형질 관련 EST profiling과 이를 이용한 EST-SSR 및 SNP 마커 개발)

  • Oh, Gyu-Dong;Hwang, Eun-Mi;Shim, Eun-Jo;Jeon, Sang-Jin;Park, Young-Doo
    • Horticultural Science & Technology
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    • v.28 no.6
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    • pp.1025-1038
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    • 2010
  • Carrot ($Daucus$ $carota$ L. var. $sativa$) is one of the most widely used crops in the world. Moreover it is an important crop because of its high content of ${\beta}$-carotene, well-known as the precursor of vitamin A carotenoid. However, seed-hair which is generated in epidermal cell of seeds inhibits absorption and germination. For that reason, carrot seeds are commercialized after mechanical hair removal process. To overcome such cumbersome weaknesses, new breeding program for developing hairless-seed carrot cultivar has been needed. Therefore, in this study, cDNA libraries from seeds of short-hair seed phenotype CT-ATR615 OP 666-13line and hairy seed CT-ATR615 OP-CK1-9 line were constructed and expression patterns related to generation of seed-hair were analyzed by comparison of EST sequences. Differential EST sequence results between two lines were classified into FunCat functional categories based on the results of BlastX search. Higher expression quantities belonging to metabolic category were shown on short-hair seed line than hairy-seed one. Differential expression quantities between those two lines in the protein folding and stabilization, subcellular localization categories were supposed to contribute variously on the generation of seed-hair. We confirmed 50 and 59 SSR sites, and 2 SNP sites by analyzing EST sequences in two lines; thereafter, we designed SNP and SSR primer sets from these EST sequence information as a molecular marker. These markers are thought to be used in research of molecular markers for classification of carrot family and related to various traits, as well as seed-hair characteristic.

Sequencing analysis of the OFC1 gene on the nonsyndromic cleft lip and palate patient in Korean (한국인 비증후군성 구순구개열 환자의 OFC1 유전자의 서열 분석)

  • Kim, Sung-Sik;Son, Woo-Sung
    • The korean journal of orthodontics
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    • v.33 no.3 s.98
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    • pp.185-197
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    • 2003
  • This study was performed to identify the characteristics of the OFC1 gene (locus: chromosome 6p24.3) in Korean patients, which is assumed to be the major gene behind the nonsyndromic cleft lip and palate. The sample consisted of 80 subjects: 40 nonsyndromic cleft lip and palate patients (proband, 20 males and females, mean age 14.2 years); and 40 normal adults (20 males and 20 females, mean age 25.6 years). Using PCR-based assay, the OFC1 gene was amplified, sequenced, and then searched for similar protein structures. Results were as follows: 1. The OFC1 gene contains the microsatellite marker 'CA' repeats. The number of the reference 'CA' repeats was 21 times, and formed as TA(CA)11TA(CA)10. But, in Koreans, the number of tandem 'CA' repeats was varied from 17 to 26 except 18, and 'CA' repeats consisted of TA(CA)n. 2. Nine allelic variants were found. Distribution of the OFC1 allele was similar between the patients and control group. 3. There was a replacement of the base 'T' to 'C' after 11 tandem 'CA' repeats in Koreans compared with Weissenbach's report. However, the difference did not seem to be the ORF prediction results between Koreans and Weissenbach's report. 4. The BLAST search results showed the Telomerase reverse transcriptase (TERT) and the Nucleotide binding protein 2 (NBP2) as similar proteins. The TERT was a protein product by the hTERT gene in the locus 5p15.33 (NCBI Genome Annotation; NT023089) The NBP2 was a protein product by the ABCC3 (ATP-binding cassette, sub-family C) gene in the locus 17q22 (NCBI Genome Annotation; NT010783). 5. In the Pedant-Pro database analysis, the predictable protein structure of the OFC1 gene had at least one transmembrane region and one non-globular region.

Expression Pattern of Skeletal-Muscle Protein Genes and Cloning of Parvalbumin mRNA in Dark-banded Rockfish (Sebastes inermis) (볼락(Sebastes inermis) 근육단백질 유전자의 성장단계별 발현 양상과 parvalbumin 유전자 클로닝)

  • Jang, Yo-Soon
    • Korean Journal of Ichthyology
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    • v.23 no.1
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    • pp.1-9
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    • 2011
  • Differentially Expressed Gene (DEG) was obtained from Differential Display Reverse Transcription (DDRT)-PCR using Annealing Control Primer (ACP) to search and clone genes related to developmental stages of Sebastes inermis. By using 120 ACPs, the nucleotide sequences obtained from 16 DEGs showing higher expression in 6-month-old skeletal muscle than 18-month-old ones and from 22 DEGs displaying stronger expression in 18-month-old than 6-month-old were analyzed and BLAST was conducted. The results identified that DEGs shared 69~95% homology with genes of parvalbumin (PVALB), nucleoside diphosphate kinase (NDK) B, tropomyosin (TPM), troponin I (TnI), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), muscle-type creatine kinase (CKM2), small EDRK-rich factor 2 (SERF2), adenosine monophosphate deaminase (AMPD), Trimeric intracellular cation channel type A (TRICA), Rho GTPase-activating protein 15 (ARHGAP15), S-formylglutathione hydrolase (Esterase D; ESD), heat shock protein 70 (hsp70), type 1 collagen alpha 2 (COL1A2), glutathione S-transferase, Mid1-interacting protein 1 (Mid1lip1), myosin light chain 1 (MYL1), sarcoplasmic/endoplasmic reticulum calcium ATPase 1B (SERCA1B), and ferritin heavy subunit (FTH1). Expression pattern by developmental stage of DEG14 and PVALB exhibiting strong expression in 6-month-old skeletal muscle was investigated using real time PCR. Expression was reduced as Sebastes inermis grew. Expression of PVALB gene was extremely low after 6 months of age. Expression of CKM2 showed higher expression in 18-month-old skeletal muscle than in 6-month-old muscles, and increased continuously until 4 years old, after which CKM2 expression became gradually reduced. By analysis of tissue-specific expression patterns of DEG, DEG14 was expressed mainly in skeletal muscle, liver, kidney and spleen tissues, whereas PVALB expression was expressed in skeletal muscle and kidney, but not in liver and spleen tissues. CKM2 was expressed in skeletal muscle, kidney, and spleen tissues, but not in liver tissues. PVALB gene was composed of 110 amino acids, which constituted 659 bp nucleotides. The results reported here demonstrate that the expression patterns of parvalbumin and CKM2 could be used as molecular markers for selecting fishes exhibiting fast growth.

The Differentially Expressed Genes by Radiotherapy in the Patients with Uterine Cervix Cancer (자궁경부암 환자에서 방사선치료 시 발현되는 유전자의 규명)

  • Seo Eun Young;Cho Moon-June;Lee Jeung Hoon;Lee Young-Sook;Na Myung-Hoon;Lee Woong-Hee;Kim Jun-Sang;Kim Jae-Sung
    • Radiation Oncology Journal
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    • v.19 no.4
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    • pp.389-396
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    • 2001
  • Purpose : To detect differentially expressed genes in the patients with uterine cervical cancer during the radiation therapy. Materials and Methods : In patients with biopsy proven uterine cervical cancer, we took tumor tissue just before radiation therapy and at 40 minutes after external irradiation of 1.8 Gy. Total RNAs isolated from non-irradiated and irradiated tumor tissue samples were analyzed using the differential-display reverse transcription-polymerase chain reaction (DDRT-PCR). Complementary DNA (cDNA) fragments corresponding to differentially expressed messenger RNAs(mRNAs) were eluted, and cloned. The differential expression of the corresponding mRNAs was confirmed by reverse northern blot. Differentially expressed cDNA bands were sequenced. Nucleotide sequence data were analyzed in the Gene Bank and EMBL databases via the BLAST network sewer to identify homologies to known genes or cDNA fragments. Expression pattern of down-regulated clone was examined using RT-PCR in S patients undergoing radiotherapy. Results : We identified 18 differentially expressed bands by DDRT-PCR, which were eluted and cloned. There were 10 up-regulated clones and 1 down-regulated clone in reverse northern blot. One cDNA fragment had homology to chemokine receptor CXCR4, four were identified as Human ESTs in the EMBL database in EST clones. Down-regulated CxCa-11 was also down regulated in all patients. Conclusion : Using the DDRT-PCR, we have identified 10 up-regulated and 1 down-regulated clone(s) in the patients with uterine cervical cancer during the radiation therapy. The clinical relevance and the functions of these genes will be further investigated.

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Isolation from Chungkookjang and Characterization of a Bacterium Producing an Extracellular Protease of High Specific Activity (청국장으로부터 고 비활성 세포외 Protease 생산 세균의 분리 및 동정)

  • Park, Hee-Jin;Park, Heui-Dong
    • Food Science and Preservation
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    • v.17 no.3
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    • pp.410-417
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    • 2010
  • Several extracellular protease-producing bacteria were isolated from Chungkookjang, a traditional Korean food of fermented soybeans, on skim milk agar plates. Among these bacteria, strain D14 exhibited the highest production (15.2 U/mL) and specific activity (40.0 U/mg protein) of extracellular protease activity as assessed on growth in a protease induction medium composed of 1% (w/v) soluble starch, 1.5% (w/v) skim milk, 0.5% (w/v) yeast extract, and 2% (w/v) NaCl. The bacterium was identified as Bacillus subtilis based on morphological and physiological characteristics and 16S rDNA sequence. A BLAST search of 16S rDNA sequences revealed that the isolate was most closely related to Bacillus subtilis subsp. subtilis strain NCIB 3610. The 16S rDNA sequence homology was 99.9%. Our isolate produced the highest level of protease when grown in a protease induction medium containing 1% (w/v) sorbitol and 0.5% (w/v) yeast extract. Fructose and glucose reduced enzyme production to 12.7% and 35.9%, respectively, of the level seen when the strain was grown in medium containing soluble starch. Soytone also reduced enzyme production to 61.4% of the level noted when the strain was grown in medium containing yeast extract.

Optimization of artificial cultivation of Tremella fuciformis in closed culture bottle (흰목이버섯 대량생산을 위한 용기내 재배 최적화 연구)

  • Choi, Sung Woo;Chang, Hyun-You;Yoon, Jeong Weon;Lee, Chan
    • Journal of Mushroom
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    • v.6 no.1
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    • pp.20-26
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    • 2008
  • The stromatal forms of T. fuciformis and the mycelia of Hypoxylon sp. were collected. The DNA sequence in the ITS region of the 5.8S ribosomal genes of isolated strain KG103 was very similar to that of T. fuciformis AF042409 with a homology of over 98% in the EMBL/GenBank database through BLAST searching. A second isolate, No KG201, one of the symbiotic strains for cultivating T. fuciformis also exhibited high homology with Annulohhypoxylon stygium AJ390406. Potato Dextrose Medium exhibited the best mycelial growth of 14 mm/14 days and 85 mm/14 days for T. fuciformis and its symbiotic fungi, respectively. Optimum culture conditions for the micelial growth were pH 5 at $25^{\circ}C$. For the optimization of artificial cultivation of T. fuciformis in bottle with sawdust medium, several conditions such as type of sawdust, supplements, pH, moisture content, and incubation temperature were investigated. T. fuciformis and symbiotic fungi showed fast mycelial growth on corn cob media (77 and 52%) followed by oak tree sawdust and cotton seed meal. The optimal temperature for mycelial growth of T. fuciformis and symbiotic fungi on corn cob media was $25^{\circ}C$ at 55% of moisture content.

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Intestine Bacterial Microbiota of Asian Hornet (Vespa Velutina Nigrithorax) and Honey Bee (등검은말벌과 꿀벌의 장내 세균 군집 비교)

  • Kim, Euyeon;Seo, Jeongwon;Yang, So Hee;Kim, In-Seon;Koo, Yeonjong
    • Korean Journal of Environmental Agriculture
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    • v.37 no.2
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    • pp.135-140
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    • 2018
  • BACKGROUND: The Asian hornet (Vespa velutina nigrithorax), a wasp species, has attacked honey bee populations and affected the beekeeping industry in Korea over the past 15 years. However, little research has been done with this invasive species. In this study, we investigated the intestine bacterial microbiota of Asian hornets and honey bees to design an attractive trap for Asian hornets. METHODS AND RESULTS: Genomic DNAs isolated from the intestine microorganisms of Asian hornets and honey bees were utilized to amplify bacterial 16S rDNA for the comparative sequence analysis. The next generation sequencing analysis identified that the orders Flavobacteriales as the most abundant intestinal microorganisms in Asian hornets, showing a clear difference compared to honey bees in which Aeromonadales are dominant. We also report five newly identified 16S rDNA sequences of Asian hornet intestinal bacteria. According to the sequence blast search, these five bacteria belong to the genera Thalassomonas, Caedobacter, Vampirovibrio, Alkaliphilus and Calothrix. CONCLUSION: While Asian hornets and honey bees show similar intestine bacterial diversity, the relative ratio of bacterial populations is different. providing useful information to design pest control agents specifically targeting Asian hornets.

Isolation and Phylogenetic Analysis of $Botryococcus$ $braunii$ (Trebouxiophyceae) from Korean Freshwaters (한국산 $Botryococcus$ $braunii$ (Trebouxiophyceae)의 분리 및 계통분류학적 분석)

  • Lee, Seung-Hoon;Lee, Chan-Hee;Jo, Beom-Ho;Ahn, Chi-Yong;Kim, Han-Soon;Oh, Hee-Mock
    • Korean Journal of Environmental Biology
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    • v.30 no.1
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    • pp.31-38
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    • 2012
  • Recently, energy security is one of the most important world-wide issues. Biodiesel derived from microalgae has received much attention as a renewable bioenergy. The green colonial alga, $Botryococcus$ $braunii$, is characterized by the ability to produce and accumulate large amounts of hydrocarbons and fatty acids. In this study, we have isolated 5 strains of $B.$ $braunii$ from Korean surface waters using a microcapillary-pipetting method and identified them by morphological features and phylogenetic analysis of the 18S rRNA gene. Phylogenetic analysis indicates that 5 strains of $B.$ $braunii$ are placed in the class of Trebouxiophyceae, and strains belong to race A type producing hydrocarbons which are alkadienes and alkatrienes. In addition, we need further studies to find out optimal growth conditions for producing biodiesel.

De novo Genome Assembly and Single Nucleotide Variations for Soybean Mosaic Virus Using Soybean Seed Transcriptome Data

  • Jo, Yeonhwa;Choi, Hoseong;Bae, Miah;Kim, Sang-Min;Kim, Sun-Lim;Lee, Bong Choon;Cho, Won Kyong;Kim, Kook-Hyung
    • The Plant Pathology Journal
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    • v.33 no.5
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    • pp.478-487
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    • 2017
  • Soybean is the most important legume crop in the world. Several diseases in soybean lead to serious yield losses in major soybean-producing countries. Moreover, soybean can be infected by diverse viruses. Recently, we carried out a large-scale screening to identify viruses infecting soybean using available soybean transcriptome data. Of the screened transcriptomes, a soybean transcriptome for soybean seed development analysis contains several virus-associated sequences. In this study, we identified five viruses, including soybean mosaic virus (SMV), infecting soybean by de novo transcriptome assembly followed by blast search. We assembled a nearly complete consensus genome sequence of SMV China using transcriptome data. Based on phylogenetic analysis, the consensus genome sequence of SMV China was closely related to SMV isolates from South Korea. We examined single nucleotide variations (SNVs) for SMVs in the soybean seed transcriptome revealing 780 SNVs, which were evenly distributed on the SMV genome. Four SNVs, C-U, U-C, A-G, and G-A, were frequently identified. This result demonstrated the quasispecies variation of the SMV genome. Taken together, this study carried out bioinformatics analyses to identify viruses using soybean transcriptome data. In addition, we demonstrated the application of soybean transcriptome data for virus genome assembly and SNV analysis.

Occurrence of Bunch Rot Disease Caused by Aspergillus tubingensis on Shine Muscat Grape (Aspergillus tubingensis에 의한 샤인머스켓 포도송이썩음병(가칭)의 발생)

  • Kim, Young Soo;Kwon, Hyeok Tae;Hong, Seung-Beom;Jeon, Yongho
    • Research in Plant Disease
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    • v.25 no.4
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    • pp.220-225
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    • 2019
  • During the year 2018, the symptoms of bunch rot on Shine Muscat (Vitis vinifera L.) were observed in Kimcheon-si, Gyeongbuk province in Korea. The disease appears on the Shine Muscat as a black rot due to prolific fungal sporulation after it has invaded into the Shine Muscat which look completely empty and dryness. Colonies of these fungi are present on the Shine Muscat skin from fruit setting and increase in amount from early season to harvest, while become peak at ripening stage. To isolate the causal agent, small fragments (2 to 3 mm) of decayed tissue from the lesion margin were placed onto potato dextrose agar (PDA) plates. Fungal colonies on PDA produced dense white aerial mycelium and then covered with dark black conidial heads. These heads were large and radiate, and vesicles were globose (2.12-32.0×2.0-3.1 ㎛). Based on morphological and cultural characteristics, this fungus was identified as Aspergillus tubingensis. To confirm its identity, the internal transcribed spacer, β-tubulin, and RNA polymerase II was sequenced for molecular identification. BLAST search indicated 99% identity with A. tubingensis. The pathogenicity test on healthy grape of Shine Muscat produced bunch rot, as the original symptoms. To select effective fungicides for the control of brunch rot, an in vitro antifungal activity of seven fungicides were evaluated against the growth of A. tubingensis. Five fungicides (dipenoconazole, tebuconazole, metconazole, iminoctadine, and captan) exhibited significantly strong suppression of the mycelial growth of A. tubingensis.