• Title/Summary/Keyword: EcoR I

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BACTERIAL IDENTIFICATION WITH RANDOM-CLONED RESTRICTION FRAGMENT OF Porphyromonas endodontalis ATCC 35406 GENOMIC DNA (무작위로 클로닝한 Porphyromonas endodontalis ATCC 35406 지놈 DNA의 제한절편 hybridization법에 의한 세균동정)

  • Um, Won-Seok;Han, Yoon-Soo
    • Restorative Dentistry and Endodontics
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    • v.20 no.2
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    • pp.645-654
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    • 1995
  • Porphyromonas endodontalis is a black-pigmented anaerobic Gram negative rod which is associated with endodontal infections. It has been isolated from infected dental root canals and submucous abscesses of endodontal origin. DNA probe is an available alternative, offering the direct detection of a specific microorganism. Nucleic-acid probes can be off different types: whole different: whole-genomic, cloned or oligonucleotide probes. Wholegenomic probes are the most sensitive because the entire genome is used for possible hybridization sites. However, as genetically similar species of bacteria are likely to be present in specimences, cross-reactions need to be considered. Cloned probes are isolated sequences of DNA that do not show cross-reactivity and are produced in quantity by cloning in a plasmid vector. Cloned probes can approach the sensitivity found with whole-genomic probes while avoiding known cross-reacting species. Porphyromonas endodontalis ATCC 35406 (serotype $O_1K_1$) was selected in this experiment to develop specific cloned DNA probes. EcoR I-digested genomic DNA fragments of P. endodontalis ATCC 35406 were cloned into pUC18 plasmid vector. From the E. coli transformed with the recombinant plasmid 4 clones were selected to be tested as specific DNA probes. Restriction-digested whole-genomic DNAs prepared from P. gingivalis 38(serotype a), W50(serotype b), A7A1-28(serotype C), P. intermedia 9336(serotype b), G8-9K-3(serotype C), P. endodontalis ATCC 35406(serotype $O_1K_1$), A. a Y4(serotype b), 75(serotype a), 67(serotype c), were each seperated on agarose gel electrophoresis, blotted on nylon membranes, and were hybridized with digoxigenin-dUTP labeled probe. The results were as follows: 1. Three clones of 1.6kb(probe e), 1.6kb(probe f), and 0.9kb(probe h) in size, were obtained. These clones were identified to be a part of the genomic DNA of P. endodontalis ATCC 35406 judging from their specific hybridization to the genomic DNA fragments of their own size on Southern blot. 2. The clones of 4.9kb(probe i) was identified to be a part of the genomic DNA of P. endodontalis ATCC 35406. but not to specific for itself. It was hybridized to P. gingivalis A7A1-28, P. intermedia G89K-3.

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Seasonal Assessment of Groundwater-Dependent Ecosystem Using Monitoring of Benthic Macroinvertebrates in Wetland (계절에 따른 습지 내 저서성대형무척추동물 모니터링을 통한 지하수의존생태계 특성 평가)

  • Jeong, Chanyoung;Choi, Ji-Woong;Moon, Hee Sun;Kim, Dong-Hun;Moon, Sang-Ho;O, Yong-Hwa;Han, Ji Yeon;Oh, Seolran;Kim, Yongcheol
    • Journal of Soil and Groundwater Environment
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    • v.26 no.6
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    • pp.130-143
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    • 2021
  • Wetlands are one of the most representative groundwater dependent ecosystems(GDEs) that require access to groundwater on a permanent or intermittent basis to maintain their biological communities and ecological processes. In this study, the seasonal characteristics of the GDEs in Baekseok Reservoir Wetland were evaluated through the monitoring of the temporal and spatial community of benthic macroinvertebrates in the wetland. The appearance of benthic macroinvertebrates appearance was changed seasonally depending on environmental factors such temperature, precipitation and water level for their habitat and it also showed the clear spatial difference in the wetland. The scores of Diversity index(H'), Richness Index (R1) and the Ecological score of benthic macroinvertebrates (TESB/AESB) were relatively high at St.3 and 4(i.e., north area) where groundwater inflows into wetland(i.e., high 222Rn conc.). The statistical analysis (ANOVA test and PCA) investigated the correlation among the benthic macroinvertebrates' community, groundwater level, wetland water level and water quality. The results showed that the community of benthic macroinvertebrates at St. 3 and 4 in Baekseok Reservoir Wetlands was spatially dependent on groundwater level and groundwater inflow. The characterization and assessment of GDEs requires understanding the hydrological, biogeochemical and biological process and this study will provide information for characterization and assessment of GDEs.

RNA Interference of Chitinase Gene in Spodoptera litura (담배거세미나방(Spodoptera litura) Chitinase gene의 RNA interference)

  • Jeon, Mi Jin;Seo, Mi Ja;Youn, Young Nam;Yu, Yong Man
    • The Korean Journal of Pesticide Science
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    • v.18 no.3
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    • pp.202-209
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    • 2014
  • RNA interference (RNAi) is the method which controls phenotypes of gene in live cells. Chitinase is the enzyme helping digestion and absorption of old cuticles during the ecdysis of insects. In order to investigate molting-inhibition effect with the chitinase related gene in Spodoptera litura, RNA was extracted from the $5^{th}$ instars. cDNA was synthesized and then we obtained about 700 bp size chitinase. After PCR products were cloned into a pGEM T-easy vector, colonies were picked. DNA was extracted from the colony cultures. EcoR I enzyme was used to check whether PCR products were inserted or not. And then we confirmed vector band of about 3 kb and insert band of about 700 bp. To synthesize the dsRNA, each DNA was cut with Spe I and Nco I enzymes (Circular DNA became lineared DNA). After synthesis of dsRNA, approximately 5 ul dsRNA was injected into the $3^{rd}$ abdominal segment of S. litura $4^{th}$ larvae. The concentration of dsRNA was about $10{\mu}g/{\mu}l$. We confirmed larval-larval molting : there were phenotypically abnormal individuals - for instance malformation, molting inhibition and change of integument color. Pupaadult molting : there were phenotypically abnormal individuals - for instance molting inhibition, change of wings and malformation. Also we could investigate the pupation, emergence and variation about noninjection, treated with DW and dsRNA. Each pupation was non-injection 83.3%, DW 78.3% and dsRNA 66.7%. Each emergence was non-injection 90.0%, DW 72.3% and dsRNA 65.0%. So we considered that chitinase dsRNA induced molting inhibition effect. But each variation was non-injection 8.9%, DW 2.9% and dsRNA 19.2%. Therefore dsRNA group showed the highest variation value. When 18 hours after injecting dsRNA, we could obtain abnormal individual.

Estimating the Carrying Capacity of a Coastal Bay for Oyster Culture -I . Estimating a Food Supply to Oysters Using an Eco-hydrodynamic Model in Geoie-Hansan Bay- (굴 양식수역의 환경용량 산정 -I. 생태계 모델을 이용한 거제 · 한산만 굴 먹이 공급량 추정)

  • Park Jong Soo;Kim Hyung Chul;Choi Woo Jeung;Lee Won Chan;Park Chung Kil
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.35 no.4
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    • pp.395-407
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    • 2002
  • A 3D hydrodynamic-ecological coupled model was applied to estimate a food supply to oysters in Geoje-Hansan Bay where is one of the oyster culturing sites in Korea, In this study, the primary productivity (PP) was adopted as an index of food supply, and the spatial patterns of average chlorophyll a concentration during a culturing seasons from September to May of the following year were simulated by the model, The numerical result showed that PP was high in the inner part of the bay and the adjacent areas of Hwado island, but low in the outer. This result indicates that PP is essentially influenced by anthropogenic nutrient loadings in the system. The model was calibrated using the field data in May which is non culturing season of oysters and a simulated phytoplankton biomass agreed fairly well with the observed data ($R^{2}=0.70$, $RE=10.3\%$). The computed food supply varied from 0.19 to $1.27\;gC/m^{2}/day$ with a mean value of $0.62 gC/m^{2}/day$ from September to May. The highest value was showed in May ($1.27 gC/m^{2}/day$) and the lowest was in February ($0.19 gC/m^{2}/day$).