• 제목/요약/키워드: strain selection

검색결과 389건 처리시간 0.027초

Highly efficient production of transgenic Scoparia dulcis L. mediated by Agrobacterium tumefaciens: plant regeneration via shoot organogenesis

  • Aileni, Mahender;Abbagani, Sadanandam;Zhang, Peng
    • Plant Biotechnology Reports
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    • 제5권2호
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    • pp.147-156
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    • 2011
  • Efficient Agrobacterium-mediated genetic transformation of Scoparia dulcis L. was developed using Agrobacterium tumefaciens strain LBA4404 harboring the binary vector pCAMBIA1301 with ${\beta}$-glucuronidase (GUS) (uidA) and hygromycin phosphotransferase (hpt) genes. Two-day precultured leaf segments of in vitro shoot culture were found to be suitable for cocultivation with the Agrobacterium strain, and acetosyringone was able to promote the transformation process. After selection on shoot organogenesis medium with appropriate concentrations of hygromycin and carbenicillin, adventitious shoots were developed on elongation medium by twice subculturing under the same selection scheme. The elongated hygromycin-resistant shoots were subsequently rooted on the MS medium supplemented with $1mg\;l^{-1}$ indole-3-butyric acid and $15mg\;l^{-1}$ hygromycin. Successful transformation was confirmed by PCR analysis using uidA- and hpt-specific primers and monitored by histochemical assay for ${\beta}$-GUS activity during shoot organogenesis. Integration of hpt gene into the genome of transgenic plants was also verified by Southern blot analysis. High transformation efficiency at a rate of 54.6% with an average of $3.9{\pm}0.39$ transgenic plantlets per explant was achieved in the present transformation system. It took only 2-3 months from seed germination to positive transformants transplanted to soil. Therefore, an efficient and fast genetic transformation system was developed for S. dulcis using an Agrobacterium-mediated approach and plant regeneration via shoot organogenesis, which provides a useful platform for future genetic engineering studies in this medicinally important plant.

이타콘산 고생산성 Aspergillus terreus 변이주의 신속 선별을 위한 효율적인 균주 스크리닝 전략 개발 (Development of an Efficient Screening Strategy for Rapid Selection of High-yielding Mutants of Itaconic Acid Biosynthesized by Fungal Cells of Aspergillus terreus)

  • 신우식;김평현;이도훈;김상용;정용섭;전계택
    • KSBB Journal
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    • 제26권3호
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    • pp.229-236
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    • 2011
  • An efficient screening method was developed for rapid selection of a few overproducers of itaconic acid (IA) among the great many mutants derived from mother strains of Aspergillus terreus. For this purpose, an attempt was made to reveal the relationships of the growth rate and sporulation of each mutant on PDA solid medium with its IA productivity in the final liquid production-culture. As a result, it was possible to classify the mutated strains into 5 groups (from [A] to [E] group) according to theirmorphologies (i.e., growth rate and sporulation extent) on the PDA slants. Notably, most of the high-yielding mutants of IA were observed to belong to [A]group which had the properties of the highest growth rate and sporulation among the 5 groups, whereas the mutant groups of [C], [D] and [E] with the contrasting morphological features showed significant reductions in their IA productivities. From these results, it was concluded that the probability of selecting IA overproducing mutants could be remarkably enhanced when the mutated colonies showing faster growth rates are firstly selected on the PDA plate, and then further screening process is performed on the basis of the sporulation extents of the mutants selected. Consequently, through the application of the strategy developed in this study, costs and time involvedin the labor-intensive task of strain improvement could be reduced to a great extent, because the time-consuming liquid culture processes did not need to performed for the unfavorable mutants belonging to the groups other than group [A].

대장균 배양 중 phe W$^+$-pheS-$^{-ts}$ System에 의한 재조합 trp$^+$ 플라스미드의 안정적 유지 (Stable Maintenance of Recombinant Plasmid Containing trp $^+$ Operon in E. coli Cultures by the phe W$^+$ -pheS$^{t8}$ System)

  • 강충민;최장원;이세영
    • 한국미생물·생명공학회지
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    • 제18권1호
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    • pp.89-93
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    • 1990
  • 재조합 pBR322-trp$^+$ 플라스미드의 숙주내 안정적 유지를 목적으로 tRNA phe 의 구조유전자인 pheW$^+$ 유전자를 pBR322-trp$^+$의 플라스미드에 도입시키고, 숙주세포로는 트립토판 생산을 위한 정상숙주 LC901의 phenylalanyl tRNA synthetase 온도감수성 변이체인 LC901-pheS-ts를 구성하여 이 온도감수성 숙주의 제한온도 (restrictive temperature)에서 재조합 trp$^+$ 플라스미드의 안정적 유지와 trp$^+$ 유전자가 미치는 효과를 조사하였다.

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Analysis of the axle load of an agricultural tractor during plow tillage and harrowing

  • Hong, Soon-Jung;Park, Seung-Je;Kim, Wan-Soo;Kim, Yong-Joo;Park, Seong-un
    • 농업과학연구
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    • 제43권4호
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    • pp.665-669
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    • 2016
  • Analysis of the load on the tractor during field operations is critical for the optimal design of the tractor. The purpose of this study was to do a load analysis of an agricultural tractor during plowing and harrowing. First, a load measurement system was developed and installed in a 71 kW agricultural tractor. Strain-gauges with a telemetry system were installed in the shaft located between the axles and the wheels, and used to measure the torque of the four driving axles. Second, field experiments were conducted for two types of field operations (plowing, harrowing), each at two gear levels (M2, M3). Third, load analysis was conducted according to field operation and gear level. At M2 gear selection for plowing, the maximum, minimum, and average (S. D.) torque values were 13,141 Nm; 4,381 Nm; and 6,971 Nm (${\pm}397.8Nm$, respectively). For harrowing, at M2 gear selection, torque values were, 14,504 Nm; 1,963 Nm; and 6,774 Nm (${\pm}459.4Nm$, respectively). At M3 gear selection for plowing, the maximum, minimum, and average (S. D.) torque values were,17,098 Nm; 6,275 Nm; and 8,509 Nm (${\pm}462.4Nm$, respectively). For harrowing at M3 gear selection, maximum, minimum, and average (S. D.) torque values were, 20,266 Nm; 2,745 Nm; and 9,968 Nm (${\pm}493.2$). The working speed of the tractor increased by approximately 143% when shifted from M2 (7.2 km/h) to M3 (10.3 km/h); while during plow tillage and harrowing, the load of the tractor increased approximately 1.2 times and 1.5 times, respectively.

Induction of pathogenicity mutants from Elsinoe fawcettii, the causal fungus of citrus scab by genetic transformation

  • Hyun, Jae-Wook;Lee, Seung-Chan;Kim, Dong-Hwan;Kim, Kwang-Sik;Lim, Han-Chul
    • 한국식물병리학회:학술대회논문집
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    • 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
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    • pp.68.2-69
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    • 2003
  • Genetic transformation carried out to induce the pathogenicity mutants from the two isolates, Elsinoe fawcettii R-34 and MUD of citrus scab fungus to hygromycin resistant by transferring plasmides (pUCATPH) that contain hygB gene. We produced protoplast for transformation by using of combinations of available enzymes including ${\beta}$-D-glucanase, ${\beta}$ -glucuronidase, Iyticase and driselase. The protoplasts regenerated at 64 $\mu\textrm{g}$/ml of hygromycin B but not 128 $\mu\textrm{g}$ in sensitivity test to identify the concentration of useful marker for the selection of transformants. Approximately 1200 and 67 hygromycin resistant isolates from strain R-34 and strain MUD, respectively, were isolated on PDA added with 200 $\mu\textrm{g}$ /ml of hygromycun B. Fifty seven and 4 of hygromycin resistant isolates from strain R-34 and MUD, respectively, did not produce necrotic lesions on the leaf in detached-leaf assay. Finally, 9 isolates were isolated from strain R-34, and these Isolates produced non or very few symptoms on seedlings of citrus in greenhouse pathogenicity test. And it's very interesting that some isolates produced melanose-like symptom on very young leaves which it was not typical symptom and somtimes produced on only expanded leaf.

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에리스리톨 고생산성 변이주인 Moniliella suaveolens var. nigra의 선별과 배양특성 (Selection and Characterization of a High Erythritol Producing Mutant of Moniliella suaveolens var. nigra)

  • 박홍우;이금숙
    • KSBB Journal
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    • 제17권3호
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    • pp.290-294
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    • 2002
  • 경제적으로 에리스리톨을 대량 생산하기 위해 Moniliella suaveolens var. nigra의 wild type을 mutation시켜 변이주를 선별하였다. 선별된 변이주는 400 g/L 포도당배지 플라스크 배양에서 에리스리톨 172 g/L, 글리세롤 20 g/L를 생산하였는데 wild-type의 배양결과와 비교하면 에리스리롤의 생산성은 비슷하고 부산물인 글리세롤의 생성은 50% 적다. 변이주는 wild type 배양시 발생하는 다량의 foam을 적게 발생시킴이 관찰되었다. 배양기 5 L 배양에서 변이주는 에리스리톨의 생산과 부산물 글리세롤의 생성이 wild-type의 것들과 비슷하였다. 이의 원인은 아직 규명되지 않았으나 배양 중 부적절한 산소전달이나 배양 중 발생하는 거품에 세포가 응집하였기 때문인 것으로 추측된다. 균주의 대사를 조사한 결과, 글리세롤은 에리스리톨로 변환되나 에리스리톨은 글리세롤로 변환되지 않음이 관찰되었다. 새로운 고생산성 Moniliella 변이주는 분리.정제 비용이 절감된 순도 놀은 에리스리톨 생성을 가능 하게 할 것이다.

인체분변으로부터 분리한 유산균 Lactobacillus pentosus Miny-148의 생균제 특성 연구 (Probiotic Property of Lactobacillus pentosus Miny-148 Isolated from Human Feces)

  • 정민영;박용하;김현수;부하령;장영효
    • 미생물학회지
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    • 제45권2호
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    • pp.177-184
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    • 2009
  • 우수한 생균제를 개발하기 위하여 안전성이 알려진 유산균을 대상으로 인체의 분변으로부터 300여 균주를 분리하고 내산성, 내담즙성, 내열성, 항균력, 항암 및 항바이러스 효과를 가지는 균주들을 선발하여 생균제 특성을 나타내는지를 알아보기 위해 본 실험을 수행하였다. 인체에서 분리한 여러 균주 중 Miny-148 균주는 낮은 pH 및 높은 담즙산에 대한 내성, 열처리에 대한 열안정성을 지녀 기초적인 probiotic 특성을 가진 균주로 선발되어, Lactobacillus pentosus (99.9% 상동성)로 동정되었다. 항균력 실험에서 Escherichia coli O157:H7을 비롯한 Shigella flexneri, Bacillus anthracis, Staphylococcus aureus, E. coli, Vibrio cholerae, V. vulnificus, Salmonella typhimurium, 그리고 및 methicillin-resistant S. aureus (MRSA) 균주 8종 등 총 16종의 병원성세균을 억제하였다. 또한 Miny-148은 결장암 세포인 HT-29 cell을 억제하였을 뿐 아니라, transmissible gastroenteritis virus의 생육을 저해하여 세포변성 억제효과를 가진 우수한 probiotic 특성을 지닌 균주로 분석되었다.

고추 탄저병 방제제 Burkholderia lata CAB13001 선발 및 포장방제 효과 (Selection of Antifungal Bacteria Burkholderia lata CAB13001 for Control on Red Pepper Anthracnose and Its Control Efficacy in Field)

  • 함수상;김병련;권미경;한광섭;박인희;서경원
    • 한국유기농업학회지
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    • 제26권4호
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    • pp.649-660
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    • 2018
  • Colletotrichum acutatum에 의해 발생하는 고추탄저병을 방제하기 위해 토양 근권으로부터 분리한 미생물 중 길항효과가 있는 미생물을 선발하여 주요 작물병원균에 대한 항균활성 효과와 포장에서의 방제효과를 검정하였다. 기내 대치배양법을 이용한 길항력을 검정하였을 때, CAB13001 균주는 고추 탄저병을 일으키는 Colletotrichum acutatum을 포함한 식물 병원균 Sclerotinia cepivorum, Sclerotinia sclerotium, Botrytis cinerea에 대한 뛰어난 항균활성 효과를 보였다. 또한 풋고추를 이용한 생체 검정에서 선발균주는 고추 탄저병의 병반진전 억제효과가 우수하였으며, 포장에서의 방제가 역시 무처리 대비 82.4%로 매우 우수하였다. 이러한 결과로 볼 때 CAB13001 균주는 공기 전염을 하는 고추 탄저병의 생물적 방제제로 매우 유용할 것으로 판단된다. 한편, 본 연구에 사용된 길항미생물 CAB13001 균주는 16s rRNA gene의 계통학적 위치를 분석한 결과 Burkholderia lata로 동정하였다.

오이 탄저병원균과 잿빛곰팡이병원균에 억제 효과가 있는 방선균 선발 (Selection of Antagonistic Soil Actinomycetes Against Both Colletotrichum orbiculare and Botrytis cinerea in Cucumber Plants)

  • 곽화숙;김지원;박진우;상미경
    • 한국유기농업학회지
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    • 제29권4호
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    • pp.575-588
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    • 2021
  • 본 연구에서는 오이 지상부에 발생하는 두 가지 병원균 B. cinerea와 C. orbiculare에 항균활성을 가지는 유용한 방선균을 선발하고자 하였다. 토양에서 분리한 방선균 560 균주 중 두 병원균에 대해 동시 항균활성을 나타낸 7균주를 일차적으로 선발한 후, 배양여액의 항균활성 검정을 위한 대치배양과 오이 식물체 검정을 통해 최종적으로 S20-465 균주를 선발하였다. 이 균주는 염기서열 비교 분석을 통해 Streptomyces sp.로 동정하였다. S20-465 균주의 배양여액은 두 병원균의 균사 생장을 60%이상 억제하였다. 배양여액의 20배, 40배 희석액을 분무 처리할 경우 오이 잎에서의 잿빛곰팡이병에 의한 병반길이를 감소시켰을 뿐만 아니라 탄저병의 발병도도 대조구에 비해 50% 이상 감소시켜 생물검정에서도 병억제 효과가 있는 것을 확인하였다. 따라서 최종 선발한 S20-465는 앞으로 생물방제 미생물소재로써 활용할 수 있을 것으로 사료된다.

Biosurfactant as a microbial pesticide

  • Lee, Baek-Seok;Choi, Sung-Won;Choi, Ki-Hyun;Lee, Jae-Ho;Kim, Eun-Ki
    • 한국생물공학회:학술대회논문집
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    • 한국생물공학회 2003년도 생물공학의 동향(XII)
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    • pp.40-44
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    • 2003
  • Soil-borne infectious disease including Pythium aphanidermatum and Rhizoctonia solani causes severe damage to plants, such as cucumber. This soil-borne infectious disease was not controlled effectively by chemical pesticide. Since these diseases spread through the soil, chemical agents are usually ineffective. Instead, biological control, including antagonistic microbe can be used as a preferred control method. An efficient method was developed to select an antagonistic strain to be used as a biological control agent strain. In this new method, surface tension reduction potential of an isolate was included in the ‘decision factor’ in addition to the other factors, such as growth rate, and pathogen inhibition rate. Considering these 3 decision factors by a statistical method, an isolate from soil was selected and was identified as Bacillus sp. GB16. In the pot test, this strain showed the best performance among the isolated strains. The lowest disease incidence rate and fastest seed growth was observed when Bacillus sp. GB16 was used. Therefore this strain was considered as plant growth promoting rhizobacteria (PGPR). The action of surface tension reducing component was deduced as the enhancement of wetting, spreading, and residing of antagonistic strain in the rhizosphere. This result showed that new selection method was significantly effective in selecting the best antagonistic strain for biological control of soil-borne infectious plant pathogen. The antifungal substances against P. aphanidermatum and R. solani were partially purified from the culture filtrates of Bacillus sp. GB16. In this study, lipopeptide possessing antifungal activity was isolated from Bacillus sp. GB16 cultures by various purification procedures and was identified as a surfactin-like lipopeptide based on the Fourier transform infrared spectroscopy (FT-IR), nuclear magnetic resonance (NMR), high performance liquid chromatography mass spectroscopy (HPLC-MS), and quadrupole time-of-flight (Q-TOF) ESI-MS/MS data. The lipopeptide, named GB16-BS, completely inhibited the growth of Pythium aphanidermatum, Rhizoctonia solani, Penicillium sp., and Botrytis cineria at concentrations of 10 and 50 mg/L, respectively. A novel method to prevent the foaming and to provide oxygen was developed. During the production of surface active agent, such as lipopeptide (surfactin), large amount of foam was produced by aeration. This resulted in the carryover of cells to the outside of the fermentor, which leads to the significant loss of cells. Instead of using cell-toxic antifoaming agents, low amount of hydrogen peroxide was added. Catalase produced by cells converted hydrogen peroxide into oxygen and water. Also addition of corn oil as an oxygen vector as well as antifoaming agent was attempted. In addition, Ca-stearate, a metal soap, was added to enhance the antifoam activity of com oil. These methods could prevent the foaming significantly and maintained high dissolved oxygen in spite of lower aeration and agitation. Using these methods, high cell density, could be achieved with increased lipopeptide productivity. In conclusion to produce an effective biological control agent for soil-borne infectious disease, following strategies were attempted i) effective screening of antagonist by including surface tension as an important decision factor ii) identification of antifungal compound produced from the isolated strain iii) novel oxygenation by $H_2O_2-catalase$ with vegetable oil for antifungal lipopeptide production.

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