• Plant Biotechnology Reports
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• 제2권4호
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• pp.267-270
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• 2008

We have successfully used the low-pressure BioWare gene gun, developed for gene transfer in animal cells, for plant tissues. The BioWare device is easy to manipulate. Just 50 psi helium pressure was sufficient to transfer foreign genes into the aleurone layer and embryo of maize without causing tissue damage in the impact area. As shown by expression signals from invasive histochemical ${\beta}-glucuronidase$ (GUS) activity, the foreign reporter gene expressed well in bombarded tissues. This successful GUS-transient expression extends the application of this low-pressure gene gun from animal cells to plant tissues.

• Asian-Australasian Journal of Animal Sciences
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• 제12권1호
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• pp.9-14
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• 1999

Factors affecting gene gun-mediated expression of the human erythropoietin gene were investigated in primary cultured oviduct cells from laying hens. The human erythropoietin gene was transfected by a gene gun method at $1.25{\mu}g$ per dish, and cultured in a synthetic serum-free medium for 72 hrs. The concentration of human erythropoietin mRNA was determined by RNA : RNA solution hybridization. In experiment 1, the effect of changing the shooting pressure of DNA-coated microparticles with nitrogen gas was tested at 20 and $60kgf/cm^2$. The results showed that the erythropoietin mRNA concentration was significantly higher at 60 than $20kgf/cm^2$. In experiment 2, the effects of supplementing the medium with fetal calf serum at 10%, and raising the shooting pressure from 60 to $80kgf/cm^2$ on the cell number and erythropoietin gene expression were examined. Although supplementation with fetal calf serum significantly increased the cell numbes compared with no supplemented controls (p < 0.05), erythropoietin mRNA concentration per $10^3$ cells was not affected. Raising the shooting pressure from 60 to $80kgf/cm^2$ did not affect either of the parameters, In experiment 3, the effect of supplementing ascorbate 2-phosphate at 0.5 mM was tested. The results indicated that the ascorbate supplementation significantly increased the cell number (p < 0.05), and tended to increase erythropoietin mRNA concentration (p < 0.1). Thus, for human erythropoietin gene expression by using the gene gun method, shooting pressure with nitrogen gas should be sufficient at $60kgf/cm^2$ and supplementation with ascorbate phosphate would be useful to enhance not only the cell proliferation but also erythropoietin gene expression.

• Journal of Microbiology and Biotechnology
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• 제26권3호
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• pp.521-529
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• 2016

Extensive use of antibiotics over recent decades has led to bacterial resistance against antibiotics, including gentamicin, one of the most effective aminoglycosides. The emergence of resistance is problematic for hospitals, since gentamicin is an important broad-spectrum antibiotic for the control of bacterial pathogens in the clinic. Previous study to identify gentamicin resistance genes from environmental samples have been conducted using culture-dependent screening methods. To overcome these limitations, we employed a metagenome-based culture-independent protocol to identify gentamicin resistance genes. Through functional screening of metagenome libraries derived from soil samples, a fosmid clone was selected as it conferred strong gentamicin resistance. To identify a specific functioning gene conferring gentamicin resistance from a selected fosmid clone (35-40 kb), a shot-gun library was constructed and four shot-gun clones (2-3 kb) were selected. Further characterization of these clones revealed that they contained sequences similar to that of the RNA ligase, T4 rnlA that is known as a toxin gene. The overexpression of the rnlA-like gene in Escherichia coli increased gentamicin resistance, indicating that this toxin gene modulates this trait. The results of our metagenome library analysis suggest that the rnlA-like gene may represent a new class of gentamicin resistance genes in pathogenic bacteria. In addition, we demonstrate that the soil metagenome can provide an important resource for the identification of antibiotic resistance genes, which are valuable molecular targets in efforts to overcome antibiotic resistance.

• 한국자원식물학회지
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• 제21권4호
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• pp.249-253
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• 2008

식물의 미토콘드리아에서 eukaryotic 유전자 발현하는 양상을 관찰한다는 것은 매우 중요하게 알여져 왔다. 본 실험에서는 식물의 미토콘드리아에서 발현하는 유전자와 GFP가 미토콘드리아에 발현하는지를 구명하였다. 미토콘드리아(mt)에서 발현 하는 AtBI-1 유전자와 GFP 유전자를 35S promoter를 가진 pBin vector에 재조합한 후, 유전자총을 이용한 형질전환법으로 담배의 잎과 cotyledon에 형질전환하여 재분화 된 shoot를 얻었다. mt에서 그 유전자가 발현 되는 것을 현미경하에서 GFP가 발광하는 것으로 확인하였다. 또한 PCR분석과 Southern분석에서도 미토콘드리아에서 AtBI-1 유전자가 발현함을 확인하였다. 따라서 본 연구의 결과로 mt에 관련된 유전자를 식물의 조직에 형질전환 하여 1개 이상의 유전자가 식물의 mt에 삽입되어 그 유전자의 특성이 발현되는데 이용되어 질수 있을것이라 생각된다.

• Journal of Plant Biotechnology
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• 제29권4호
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• pp.223-228
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• 2002

본 연구실에서 개발된 particle inflow gun (PIG)은 조작이 간편하고, 사용비용도 저렴하며, 식물 세포 내의 유전자 도입효율이 높은 특징을 갖고 있다. PIG 장비를 이용하여 벼 캘러스 내로의 유전자 도입 조건을 검토하기 위해서 사용된 vector는 pIG121Hm으로서 T-DNA 내부에 intron GUS ($\beta$-glucuronidase)와 hygromycin 및 kanamycin 저항성 유전자를 포함하고 있다. 또한 벼 캘러스 내에 물리적으로 DNA를 도입할 때에 DNA 도입 효율과 관계가 높은 요인들을 GUS의 발현빈도를 통하여 조사하였다. 그 결과 gold particle에 DNA를 부착하는 과정에 사용되는 spermidine과 calcium chloride의 경우 무첨가구에 비해 16 mM의 spermidine과 1.5 M의 calcium chloride 첨가구에서 GUS 발현율이 각각 2배, 3배 증가하였다. 그리고 1회 분사되는 gold particles양이 2 mg의 경우 가장 높은 GUS 발현율을 보여주었으며, 또한 PIG장비의 분사거리와 헬륨의 압력은 벼의 배양세포의 경우 12cm의 분사거리에서 3.5 bar (50 psi)의 헬륨압력으로 분사하였을 때 GUS 발현율이 가장 높았다. 이상의 결과에서 PIG 장비를 이용한 유전자 도입은 본 연구에서 검토한 최적의 조건을 이용하였을 경우 기존에 많이 사용되고 있는 Biolistic Gun (Bio-Rad 사)과 거의 비슷한 유전자 도입효율을 보여 주었다. 특히 PIG 장비의 경우 조작이 매우 간편하고, 분사에 사용되는 일회용 부품이 필요하지 않기 때문에 대량의 반복실험을 필요로 하는 연구에서 손쉽게 사용되리라 기대된다.

• 한국동물위생학회지
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• 제46권4호
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• pp.339-348
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• 2023

The outbreak of lumpy skin disease (LSD), caused by LSD virus (LSDV), in Jeollabuk-do was first confirmed at a Korean cattle farm in Buan-gun on October 24, 2023. Afterwards, thirteen cases (twelve cases in Gochang-gun and a case in Imsil-gun) were further confirmed, resulting in a total of fourteen cases over 25 days until November 17, 2023. Clinical examination were conducted on infected and co-habitting cattle from the LSD-affected farms with particular focus on the presence of nodules throughout the body such as head, neck, chest, femur, head, and perineum. As a results, abnormal clinical signs were observed in fifteen cows: loss of appetite in six cows, high fever in three cows, eye mucosal nodules in a cow, nasal mucosal nodules in six cows, nodules on perineum in five cows, scrotum nodules in two cows, papillary nodules in a cow, and/or skin nodules in eleven cows. By the PCR methods, the common gene of capripox virus and/or the specific gene of LSDV were detected in 35 of the 69 cows tested this study. In the Farm1, capripox virus-specific gene, LSDV wild strain-specific gene, and LSDV vaccine strain-spcific gene were simultaneously detected in affected cows, indicating the cattle farm was affected by various strain of LSDV. As a result of combining clincal examination and PCR test, it was found that clinically and subclinically infeted cows coexist in the LSDV-infected farms. These finding in this study will be a great help in diagnosis and prevention of the LSD in Korean cattle farms.

• Parasites, Hosts and Diseases
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• 제49권1호
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• pp.85-90
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• 2011

Relatively little has been studied on the AMA-1 vaccine against Plasmodium vivax and on the plasmid DNA vaccine encoding P. vivax AMA-1 (PvAMA-1). In the present study, a plasmid DNA vaccine encoding AMA-1 of the reemerging Korean P. vivax has been constructed and a preliminary study was done on its cellular immunogenicity to recipient BALB/c mice. The PvAMA-1 gene was cloned and expressed in the plasmid vector UBpcAMA-1, and a protein band of approximately 56.8 kDa was obtained from the transfected COS7 cells. BALB/c mice were immunized intramuscularly or using a gene gun 4 times with the vaccine, and the proportions of splenic T-cell subsets were examined by fluorocytometry at week 2 after the last injection. The spleen cells from intramuscularly injected mice revealed no significant changes in the proportions of CD8$^+$ T-cells and CD4$^+$ T-cells. However, in mice immunized using a gene gun, significantly higher (P<0.05) proportions of CD8$^+$ cells were observed compared to UB vector-injected control mice. The results indicated that cellular immunogenicity of the plasmid DNA vaccine encoding AMA-1 of the reemerging Korean P. vivax was weak when it was injected intramuscularly; however, a promising effect was observed using the gene gun injection technique.

• 한국균학회지
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• 제46권4호
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• pp.495-503
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• 2018

2016년 4월부터 6월까지 표고 원목재배 임가 4곳과 표고 톱밥재배 임가 1곳의 재배사 내 실내 공기에서 진균을 포집 및 분리 동정하였다. 총 6개의 국내 미기록 진균(Cenangium acuum, Periconia macrospinosa, Neopestalotiopsissurinamensis, Metarhiziummarquandii, Trichoderma petersenii, Trichoderma paratroviride)을 분리하였으며 이들 균류에 대하여 형태학적 특성 및 internal transcribed spacer rDNA region, large subunitregion, ${\beta}-tubulin$ 또는 translation elongation factor $1{\alpha}$유전자 염기서열에 기반하여 계통학적 분석 결과를 기술하였다.

• 한국균학회지
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• 제45권4호
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• pp.386-393
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• 2017

우리나라 양파 재배지의 주요 곰팡이병인 양파 노균병의 원인균 Peronospora destructor를 채집하여, internal transcribed spacer (ITS) 영역을 포함한 4개 유전자의 염기서열을 분석하여 상동성 비교와 분자계통학적 유연관계를 분석하였다. 그 결과, 경남 창녕군, 함안군, 합천군과 전남 무안군, 신안군, 해남군에 발생하는 P. destructor의 4종의 유전자 염기서열 모두 100% 상동성을 보였고, 유연관계 분석에서도 모두 동일한 계통으로 확인되었다. 본 연구에서는 ITS 영역 유전자 염기서열을 이용하여 P. destructor 검출용 PCR법을 개발하였고, 노균병균만을 특이적으로 검출하였다. 또한 식물체를 포함한 total genomic DNA로 검출한계를 검정한 결과, $0.7ng/{\mu}L$까지 가능하였다. 양파 노균병균 검출용 PCR법은 육안상 병징이 나타나지 않을 때도 충분히 감염유무가 확인되었다.

• Journal of Photoscience
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• 제10권3호
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• pp.245-249
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• 2003

The psbA gene of photo system II was cloned and characterized from the P. ginseng chloroplast. The psbA gene is composed of 1,062 nucleotides. The overall amino acid sequence shows 99% and 98% identities to dicots and monocots of higher plants, respectively. Southern blot analysis revealed that a single copy of the psbA gene existed in the chloroplast genome. Northern blot analysis of the in vivo accumulation of the psbA transcript, after being grown under the different intensities (5%, 10%, 20%, and 100%) of daylight, indicated that the steady-state level of the psbA transcript was not significantly affected by light intensity.