분자농업의 현황 및 전망

김태금,양문식,Kim, Tae-Geum,Yang, Moon-Sik 한국식물생명공학회 2010 식물생명공학회지 Vol.37 No.3

Molecular farming is production of pharmaceutically and industrially important proteins in plants. Plants and plant cell culture systems have been used as bio-factory to produce recombinant proteins such as monoclonal antibodies, enzymes, vaccines, hormones, interleukins, commercial enzymes and etc. The terms molecular farming, biofarming, molecular pharming, phytomanufacturing, recombinant or plant-made industrials, planta-pharma, plant bioreactors, plant biofactory, and pharmaceutical gardening are used interchangeably. Molecular farming can provide safe and inexpensive pharmaceutical proteins as well as commercial ones. In spite of several advantages of molecular farming such as safety and inexpensive cost, there are also a couple of drawbacks in the existing technology. One of them is low expression level of target gene in plants, which has been improved by optimizing gene-based codon usage, screening of strong promoters, expression of transcription factors, subcellular targeting of target proteins, chloroplast transformation, and transient expression using viral expression system (magnifection). Some plant-based commercial proteins have already been in markets and more than twenty plant-based pharmaceuticals have been in clinical trials, from that we can expect that several plant-based pharmaceutical proteins will be seen in the markets in the near future.

형질전환 담배에서 Amaranthus 저장단백질인 AmA1 유전자의 발현

김태금,김영숙,권태호 한국식물생명공학회 2000 식물생명공학회지 Vol.27 No.3

A 1,183bp cDNA, AmA1, encoding the seed storage protein of Amaranthus hypochondriacus was isolated by reverse transcriptase-polymerase chain reaction (RT-PCR) and characterized. AmA1 gene was subcloned into plant binary vector under Cauliflower Mosaic Virus (CaMV) 35S promoter and nopaline synthase terminator (3'NOS). The recombinant binary vector was used to transform Nicotiana tabacum using Agrobacterium tumefacien -mediated transformation procedure. Shoots were induced on MS medium with 0.1 mg/L NAA, 1.0 mg/L BA, 100 mg/L kanamycin and 250 mg/L cefotaxime. Transgenic plants were selected on rooting medium based on MS medium containing 200 mg/L kanamycin and 250 mg/L cefotaxime without phytoregulators. The presence of AmA1 gene in the transgenic plants was confirmed by PCR followed by DNA hybridization. The expression of AmA1 gene in the transgenic plant was observed by RT-PCR method. Amaranthus hypochondriacus의 저장 단백질을 encoding 하는 AmA1 유전자를 RT-PCR 방법을 이용하여 분리하고 특성화하였다. AmAl 유전자를 담배에 형질전환 시키기 위해 CaMV 35S promoter와 3'NOS를 가지고 있는 식물 발현 vector에 subcloning 하고 이 재조합 vector를 이용하여 Agrobacterium-mediated형질전환 방법을 이용하여 담배에 도입시켰다 신초는 0.1 mg/L NAA, 1.0 mg/L BA, 100 mg/L kanamycin 그리고 250 mg/L cefotaxime이 첨가된 MS 선발 배지에서 선발했고, 선발된 신초는 식물 생장 조절제를 첨가 하지 않고 200 mg/L kanamycin과 250 mg/L cefotaxime이 첨가된 MS배지에서 뿌리를 유도하였다. 선발된 담배의 게놈내의 AmA1 유전자의 존재는 PCR 방법과 hybridization을 이용하여 확인되었고, AmA1 유전자의 발현은 RT-PCR방법과 Southern blot hybridization을 사용하여 확인되었다.

Production of Monoclonal Antibodies against the FimA Protein of Porphyromonas gingivalis in Nicotiana benthamiana

김태금,Bang Geul Kim,Dong-Keun Jeong,장용석,이진용,양문식 한국생물공학회 2012 Biotechnology and Bioprocess Engineering Vol.17 No.2

Porphyromonas gingivalis, a gram-negative anaerobic oral bacterium, causes periodontal disease by binding to saliva-coated oral surfaces. The FimA protein from P. gingivalis is a crucial pathogenic component of the bacterium and a target for vaccine development against periodontal disease. Complementary DNAs encoding the heavy and light chains of two monoclonal antibodies that bind specifically to the FimA protein were cloned into a plant expression vector under the control of the duplicated Cauliflower Mosaic Virus 35S promoter, and agroinfiltration was used to allow the vectors to infiltrate tobacco plants. The expressions of the heavy and light chains in the leaf tissue were detected using antibodies specific to each antibody chain. Western blot analysis showed the specific binding of the plant-derived monoclonal antibodies to the native FimA protein purified from P. gingivalis. Our finding that plant-derived monoclonal antibodies bound specifically to the native FimA protein indicates that plantderived monoclonal antibodies can protect against P. gingivalis invasion.

Immunogenicity of Fusion Protein of Cholera Toxin B Subunit- Porphyromonas gingivalis 53-kDa Minor Fimbrial Protein Produced in Nicotiana benthamiana

김태금,Tran Thuy Lan,이진용 한국생물공학회 2019 Biotechnology and Bioprocess Engineering Vol.24 No.6

Porphyromonas gingivalis induces destruction of periodontal tissues that surround and support the teeth, contributing to the development of periodontitis, which eventually results in tooth loss in adults. A 53-kDa protein of P. gingivalis is a major subunit variant protein of minor fimbriae (Mfa1), and is reported to be highly immunogenic and considered as a potential vaccine candidate. The gene encoding 53-kDa protein, was divided into three segments, and each DNA segment was fused to the gene coding for cholera toxin B subunit (CTB) to improve gut mucosal immune responses. The ctb-mfa1 fusion genes were expressed in the leaf tissues of Nicotiana benthamiana using agroinfiltration. Immunoblot analysis revealed that CTBMfa1 fusion proteins were produced in the agroinfiltrated leaves. The interaction of the plant-produced CTB-Mfa1 fusion proteins with GM1-ganglioside, which acts as the binding site for native CTB, was confirmed by GM1- ELISA. Mice immunized orally with the agroinfiltrated leaf powder containing the CTB-Mfa1 fusion proteins elicited serum IgG and fecal IgA antibodies to CTB and Mfa1. These results suggest that CTB-Mfa1 fusion proteins produced in plants can be used as an oral vaccine to control P. gingivalis infection-associated periodontitis.

분자농업의 현황 및 전망

김태금,양문식 한국식물생명공학회 2010 JOURNAL OF PLANT BIOTECHNOLOGY Vol.37 No.3

Molecular farming is production of pharmaceutically and industrially important proteins in plants. Plants and plant cell culture systems have been used as bio-factory to produce recombinant proteins such as monoclonal antibodies, enzymes, vaccines, hormones, interleukins, commercial enzymes and etc. The terms molecular farming,biofarming, molecular pharming, hytomanufacturing, recombinant or plant-made industrials, planta-pharma, plant bioreactors, plant biofactory, and pharmaceutical gardening are used interchangeably. Molecular farming can provide safe and inexpensive harmaceutical proteins as well as commercial ones. In spite of several dvantages of molecular farming such as safety and inexpensive cost, there are also a couple of drawbacks in the existing technology. One of them is low expression level of target gene in plants, which has been improved by optimizing gene-based codon usage, screening of strong promoters, expression of transcription factors, subcellular targeting of target proteins, chloroplast transformation, and transient expression using viral expression system (magnifection). Some plant-based commercial proteins have already been in markets and more than twenty plant-based pharmaceuticals have been in clinical trials, from that we can expect that several plant-based pharmaceutical proteins will be seen in the markets in the near future.

Current Trends in Edible Vaccine Development using Transgenic Plants

김태금,양문식 한국생물공학회 2010 Biotechnology and Bioprocess Engineering Vol.15 No.1

Immunology textbooks currently report orally administered antigens as inducing immune tolerance rather than immune stimulation. Nevertheless, current plant-based edible vaccine technology, if sufficiently developed, may offer several advantages. For example, it is easy to apply, store, and transport. It could also induce both mucosal and systemic immune responses, which cannot be achieved using an injection vaccine. Plant-based vaccines are also anticipated to prove quite useful in the animal industry, since the cost of injection is a significant burden in the animal industry. Although no commercial plant-based edible vaccines are currently available, several candidate vaccines are undergoing clinical trials. Consequently, many scientists are anticipating that a commercial plant-based edible vaccine will be available in the near future.

Vertical Growth of ZnO Nanorods on ITO Substrate by Using a Two-step-potential Electrochemical Deposition Method

김태금,장진탁,류혁현,이원재 한국물리학회 2013 THE JOURNAL OF THE KOREAN PHYSICAL SOCIETY Vol.63 No.1

In this study, we grew vertical ZnO nanorods on indium-doped tin-oxide (ITO) substrate by using an electrochemical deposition. For the growth, we used a two-step, continuous potential process without a template, additives, catalysts or a seed buffer layer. The morphological, structural and optical properties of ZnO were investigated using field-emission scanning electron microscopy (FESEM),X-ray diffraction (XRD) and photoluminescence (PL), respectively. The best test conditions resulted in the growth of vertical ZnO nanorods with a height of 550 nm and occurred when a first-step potential of -1.2 V was applied for 10 s, and a second-step potential of -0.7 V was applied for 1190 s. ZnO nanorods grown at the optimized growth condition show the strongest (002)peak intensity in the XRD spectrum and the highest near band edge emission over deep level edge emission (NBE/DLE) peak ratio of 76.26 and the lowest full width at half maximum (FWHM) of 144 meV in the photoluminescence spectrum.In our study, the structural and the optical properties of ZnO nanorods were highly improved,as compared with a constant single potential for 1200 s,when a first-step potential of -1.2 V was applied for 10 s and was followed by a second-step potential of -0.7 V which was applied for 1190 s.

Amaranthus 저장단백질인 AmA1유전자를 이용한 상추(Lactuca sativa)의 형질전환

김태금(Tae Geum Kim),김영숙(Young Sook Kim) 한국원예학회 2000 Horticulture, Environment, and Biotechnology Vol.41 No.5

합성유전자를 이용한 식물단백질의 향상

김태금(Tae Geum Kim),양문식(Moon Sik Yang) 한국생물공학회 1992 KSBB Journal Vol.7 No.3

Synthesis and Assembly of Escherichia coli Heat-labile Enterotoxin B Subunit in Transgenic Rice (Oryza sativa L.)

양문식,김태금,Maria Oszvald,Tae-Jin Kang,Barnabas Jenes,Laszlo Tamas 한국생물공학회 2007 Biotechnology and Bioprocess Engineering Vol.12 No.6

Escherichia coli heat-labile enterotoxin B subunit (LTB) can be used as a potent mucosal immunogen and immunoadjuvant for co-administered antigens. The synthetic LTB (sLTB) was modified based on plant optimized codon usage, and fused to a translation signal (the Kozak sequence) in the front of start codon and the ER retention signal, SEKDEL, in the c-terminus of sLTB gene. The sLTB and the wild-type LTB gene (wLTB) were located into plant expression vectors under the control of the wheat Bx17 HMW (High Molecular Weight) glutenin endosperm-specific promoter containing the first intron of the rice actin1 gene. Both genes were introduced into rice cells (Oryza sativa L.) via particle bombardment mediated transformation. The integration of LTB gene into the chromosome of transgenic plants was confirmed by genomic DNA PCR amplification methods. The transcription and translation of the LTB genes were demonstrated by reverse-transcription PCR (RT-PCR) and Western blot analyses, respectively. The LTB proteins produced in the seed tissues of transgenic rice showed binding affinity for GM1 ganglioside, a receptor for biologically active LTB, suggesting the plant-produced LTB are capable of forming active pentamers. The expression level of sLTB was higher than wLTB in transgenic rice plants and was up to 2.7% of the total soluble proteins of the seed tissues.