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Production of Human Serum Albumin in Chloroplast-Transformed Tobacco Plants
유장렬,고석민,Hyun-Chul Kim,Byung-Ho Yoo,우제욱,정화지,Dong-Woog Choi 한국식물생명공학회 2006 JOURNAL OF PLANT BIOTECHNOLOGY Vol.33 No.4
Human serum albumin (HSA) is the most abundant proteinin plasma and is the most often used intravenous protein inmany human therapies. However, HSA is currently extractedonly from plasma because commercially feasible recombinantexpression systems are not available. This studyattempted to develop an efficient system for recombinantHSA production by chloroplast transformation of tobacco. AHSA cDNA was isolated from a cDNA library constructedwith human liver tissue. Chloroplast transformation vectorswere constructed by introducing various regulatory elementsto HSA regulatory sequences. Vectors were delivered byparticle bombardment into leaf explants and chloroplasttransformedplants were subsequently regenerated intowhole plants. Southern blot analysis confirmed that the HSAcDNA was incorporated between rps12 and orf70B of thechloroplast genome as designed. Western blot analysisrevealed that hyper-expression and increasing the stabilityof HSA were achieved by modification of the regulatorysequences using the psbA5'UTRs in combination with elementsof the 14 N-terminal amino acids of the GFP and theFLAG tag. However, only plants transformed with the vectorcontaining all of these elements were able to accumulateHSA.
고등식물의 엽록체 형질전환: 원핵생물과 진핵생물의 조우
유장렬,정화지,서영배,정원중,민성란 한국식물생명공학회 2006 JOURNAL OF PLANT BIOTECHNOLOGY Vol.33 No.3
Chloroplasts are believed to be descended from certain cyanobacteria, which were taken up by phagocytosis into a host cell and lived there in a symbiotic relationship. In contrast to the current static concept on the chloroplast genome, its dynamism has been recently demonstrated: the chloroplast genome is active in intramolecular homologous recombination, producing subgenomic circles when it obtains homolgous sequences via genetic transformation. Chloroplast tranformation in higher plants provides many advantages over nuclear transformation that include higher expression levels of transgenes, polycistronic expression of transgenes, and maternal transmission of transgenes. Tobacco has been used as a model for chloroplast genetic transformation. However, it is recently possible to transform the chloroplasts of other major food and economic crops including rice, soybean, and cotton. Chloroplast-transformed crops will be able to replace bioreactors using microorganisms for production of value-added proteins in future.
유장렬,김석원,정회일 한국식물생명공학회 2006 JOURNAL OF PLANT BIOTECHNOLOGY Vol.33 No.3
Plant metabolomics is a plant biology field for identifying all of the metabolites found in a certain plant cell, tissue, organ, or whole plant in a given time and conditions and for studying changes in metabolic profiling as time goes or conditions change. Metabolomics is one of the most recently developed omics for holistic approach to biology and is a kind of systems biology. For holistic approach, metabolomics frequently uses chemometrics or multivariate statistical analysis of metabolic profillings. In plant biology, metabolomics is useful to determine functions of genes often in combination with DNA microarrays by analyzing tagged mutants of the model plants Arabidopsis and rice. This review paper attempted to introduce basic concepts of metabolomics and practical uses of multivariate statistical analysis of metabolic profiling obtained by 1H NMR and Fourier transform infrared spectrometry.
고구마에서 ADP-Glucose Pyrophosphorylase Small Subunit cDNA의 Antisense 발현에 의한 전분생합성 저해
유장렬,민성란,한지학,정원중,이영복 한국식물생명공학회 2007 JOURNAL OF PLANT BIOTECHNOLOGY Vol.34 No.4
Embryogenic calluses derived from shoot apical meristem explants of sweetpotato were subjected to particle bombardment to generate transgenic plants for antisense expression of cDNAs encoding two different AGPase small subunit (ibAGP1 and ibAGP2). Plants were generated via somatic embryogenesis. PCR and Southern analysis demonstrated that the incorporation of ibAGP1 and ibAGP2 into the genome in an antisense orientation. Immunoblot analysis confirmed reduced levels of AGPase small subunit in transgenic plant leaves. Plants with both ibAGP1 and ibAGP2 produced a lower level of the protein than plants with ibAGP1 alone. Iodine test demonstrated that transgenic plant leaves and storage root accumulated reduced amounts of starch. Iodine staining of leaf tissues indicated that transgenic plants accumulated less amount of starch than control. In accordance with western blot analysis, plants with both ibAGP1 and ibAGP2 accumulated a lower amount of starch than plants with ibAGP1 alone. Both transgenic plants exhibited a severely retarded growth, resulting in bare survival. It is suggested that disrupted expression of the gene encoding AGPase small subunit is lethal to the growth of sweetpotato contrast to other species including potato.
유장렬,고석민,인동수,정화지,최동욱 한국식물생명공학회 2007 JOURNAL OF PLANT BIOTECHNOLOGY Vol.34 No.4
This study describes conditions for the mass production of activation-tagged mutant hairy root lines of ginseng by cocultivation with Agrobacterium rhizogenes. Because it is not currently possible to produce progeny from transgenic ginseng, a loss-of-function approach for functional genomics cannot be appliable to this species. A gain-of-function approach is alternatively the choice and hairy root production by cocultivation of A. rhizogenes would be most practical to obtain a large number of mutants. Various sources of explants were subjected to genetic transformation with various strains of A. rhizogenes harboring the activation-tagging vector pKH01 to determine optimum conditions for the highest frequency of hairy root formation on explants. Petiole explants cocultivated with A. rhizogenes R1000 produced hairy roots at a frequency of 85.9% after 4 weeks of culture. Conditions for maximum growth or branching rate of hairy roots were also investigated by using various culture media. Petiole explants cultured on half strength Schenk and Hildebrandt medium produced vigorously growing branched roots at a rate of 2.6 after 4 weeks of culture. A total of 1,989 lines of hairy root mutants were established in this study. These hairy root lines will be useful to determine functions of genes for biosynthesis of ginsenosides.
적은 수의 거대 엽록체를 가진 핵 형질전환 식물체를 이용한 담배 엽록체 형질전환 빈도 제고
유장렬,정원중,민성란 한국식물생명공학회 2007 JOURNAL OF PLANT BIOTECHNOLOGY Vol.34 No.3
In the chloroplast transformation process, a chloroplast containing transformed chloroplast genome copies should be selected over wild-type chloroplasts on selection medium. It is more effective for a cell to become homoplasmic if the cell contains smaller number of chloroplasts. Therefore, to reduce the number of chloroplasts in mesophyll cells in tobacco, we overexpressed FtsZ to generate transgenic plants, of which mesophyll cell contained a few enlarged chloroplasts contrast to a wild-type mesophyll cell containing approximately 100 chloroplasts. It was demonstrated that transgenic leaf tissues comprising cells with a few enlarged chloroplasts gave rise to approximately 40% higher frequency of chloroplast-transformed adventitious shoots.