• BMB Reports
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• 제53권4호
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• pp.229-233
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• 2020

The anti-colorectal cancer monoclonal antibody CO17-1A (mAb CO), which recognizes the tumor-associated antigen EpCAM, was expressed in transgenic Arabidopsis plants. PCR and western blot analyses showed the insertion and expression of heavy chain (HC)/HC fused to the KDEL ER retention modif (HCK) and light chain (LC) of mAb CO and mAb CO with HCK (mAb COK) in Arabidopsis transformants. Both plant-derived mAb^P CO and mAb^P COK were purified from a biomass of approximately 1,000 seedlings grown in a greenhouse. In sandwich ELISA, both mAb^P CO showed a slightly higher binding affinity for the target, EpCAM, compared to mAb^M CO. In cell ELISA, both mAbs^P COs showed binding affinity to the human colorectal cancer cell line SW480. Furthermore, mAb^M CO, mAb^P CO, and mAb^P COK exhibited dose and timedependent regression effects on SW480 cells in vitro. In summation, both mAb^P CO and mAb^P COK, expressed in Arabidopsis, recognized the target antigen EpCAM and showed anti-proliferative activity against human colorectal cancer cells.

• 한국식물학회:학술대회논문집
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• 한국식물학회 1999년도 제13회 식물생명공학심포지움 New Approaches to Understand Gene Function in Plants and Application to Plant Biotechnology
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• pp.11-15
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• 1999

In order to evlauate feasibility of the gene tagging by the maize transposable element Ac in heterologous plant systems, we have investigated physical distances and directions of transposition of the element in Arabidopsis thaliana and tobacco cultured cell line BY-2. We prepared a T-DNA construct that carried a non-autonomous derivative of Ac with a site for cleavage by endonuclease I-Scel (designated dAc-I-RS element). Another cleavage site was also introduced into the T-DNA region outside dAc-I-RS. A number of transgenic Arabidopsis plants were generated, each of which had a single copy of the T-DNA at a different chromosomal location. To examine the pattern of transposition, three out of these transgenic plants were crossed with the Arabidopsis plant that carried the gene for Ac transposase and progeny in which dAc-I-RS had been transposed were isolated. After digestion of the genomic DNA of these progeny with I-SceI, sizes of segment of DNA were determined byd pulse-field gel electrophoresis. We also performed linkage analysis for the transposed elements and sites of mutations near the elements. Our results with three transgenic lines showed that 50% of all transposition events had occurred within 1,700 kilo-base pairs (kb) on the same chromosome, with 35% within 200 kb, and that the elements transposed in both directions on the chromosome with roughly equal probability. The data thus indicate that the Ac-Ds system is most useful for tagging of genes that are present within 200 kb of the chromosomal site of Ac in Arabidopsis. In addition, determination of the precise localization of the transposed dAc-I-RS element should definitely assist in map-based cloning of genes around insertion sites. In the present paper, we report typical examples of such gene isolation studies.

• 생명과학회지
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• 제12권1호
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• pp.67-76
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• 2002

Agrobacterium(LBA4404/pBI1121)을 이용하여 형질전환된 애기장대 (Arabidopsis thaliana)를 대상으로 T2, T3, F3세대에서의 도입된 외래 유전자의 비활성화 현상을 조사하였다. Kanamaycin저항성 개체들의 GUS유전자 발현을 분석한 결과, T2세대에서 조사된 12계통 중 5계통에서 GUS 비활성 개체가 관찰되었다 (GUS유전자 비활성율 2.3%). Multi copy T-DNA 계통을 조사한 결과, GUS 비활성 정도가 더욱 심해짐이 관찰되었다 (5.8%). T3 세대에서 single copy T-DNA 계통들은 1.3%의 GUS 비활성율을 보인 반면, multi-copy T-DNA 계통에서의 비활성율은 12.6%로 급격히 증가하였다. 유사한 현상이 형질전환 식물체와 정상개체를 교배하여 생산된 F2 계통에서도 관찰되었다 (비활성율 9.9%). 본 실험으로 식물체에 도입된 외래 유전자가 후대에서의 전이과정동안 점진적으로 비활성화되고, 이 현상은 multi copy T-DNA 계통에서 훨씬 심각함이 밝혀졌다.

• Journal of Plant Biology
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• 제39권4호
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• pp.249-256
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• 1996

Beet curly top virus is the DNA virus that is providing useful for basic studies of the infection of Arabidopsis thaliana with viral host and provides a system for studying both resistance and the molecular basis of symptom development. An importnat aspect of symptom development observed in BCTV-infected A. thaliana (ecotype Sei-O) was the induction of cell division on phloem and surrounding cortex cells. Analysis of the expression of GUS reporter gene activity in transgenic plants containing constructs with promoter of the auxin-inducible saur gene showed that saur promoter activity was induced concomitantly in symptomatic tissues at the inflorescence shoot tips of the transgenic lines. The auxin sensitivity tests showed that hypersusceptible ecotype, Sei-O produced more amounts of callus than susceptible ecotype, Col-O. These studies indicated that changes in auxin concentration were involved in the induction of cell division in BCTV-infected plants and clearly demonstrated that there was a strong correlation between auxin-induced gene expression and the activation of cell division.

• Journal of Microbiology and Biotechnology
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• 제24권10호
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• pp.1421-1426
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• 2014

Although a large number of AroA enzymes (EPSPS: 5-enopyruvylshikimate-3-phosphate synthase) have been identified, cloned, and tested for glyphosate resistance, only two AroA variants, derived from Agrobacterium tumefaciens strain CP4 and Zea mays, have been utilized to produce the commercial glyphosate-resistant crops. Here, we have used a PCR-based twostep DNA synthesis method to synthesize an aroA gene ($aroA_{A.\;metalliredigens}$) from Alkaliphilus metalliredigens, encoding a new EPSPS. Furthermore, transgenic Arabidopsis with the new $aroA_{A.\;metalliredigens}$ gene was obtained to confirm the potential of the novel aroA gene in developing glyphosate-resistant crops.

• 한국육종학회지
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• 제42권5호
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• pp.455-462
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• 2010

Purple acid phosphatase is important for phosphorus remobilization in plants, but its role in plant adaptation to low phosphorus availability is not known. The cDNA encoding O. sativa purple acid phosphatase (OsPAP1) has 1008 bp with an open reading frame of 335 amino acid residues. The amino acid sequence of OsPAP1 cDNA shows of 50-51% identity with other plant purple acid phosphatases. OsPAP1 was expressed in rice plants and in cell cultures in the absence of phosphate ($P_i$). The expression was organ-specific with the strongest expression in $P_i$-deprived roots. Functional expression of the OsPAP1 gene in the transgenic Arabidopsis line was confirmed by northern and western blot analysis. OsPAP1 overexpression lines had higher phosphatase activity than wild-type. Overexpression of OsPAP1 in Arabidopsis plants resulted in increased Pi accumulation under Pi sufficient condition. These results show that the OsPAP1 gene represents more efficient $P_i$ uptake and can be used to develop new transgenic dicotyledonous plants.

• Journal of Plant Biotechnology
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• 제42권4호
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• pp.409-413
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• 2015

본 연구는 토양에 오염된 중금속을 제거하기 위한 식물정화공정에 사용할 식물체를 개발하기 위해 $TgMTP_1$ 유전자를 CaMV35S 상시 발현 할 수 있도록 Ti-plasmid 벡터를 구축하여 형질전환식물을 육성하였다. 유전자가 도입된 형질전환 애기장대에서 TgMTP유전자를 과발현하는 호모 TG-116 (T3 generation) 계통을 육성하여 그 특성을 조사하였다. 호모계통으로 육성한 TG-116 계통은 callus 및 식물체에서 중금속에 대한 저항성을 보였다. 특히 RT-PCR 및 Western 분석에서 유전자의 발현은 잎에서 높게 나타났으며, Ni 흡수 및 축적이 많이 일어났다. 따라서 MTP1 유전자가 발현되어 액포에 중금속을 축적하는 실험결과를 활용한다면 식물정화공정에 사용할 수 있는 다양한 유전자원으로 기대할 수 있다.

• 한국식물생명공학회:학술대회논문집
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• 한국식물생명공학회 2005년도 추계학술대회 및 한일 식물생명공학 심포지엄
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• pp.356-360
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• 2005

Xanthine dehydrogenase (XDH), a classic enzyme involved in purine catabolism, can catalyze the formation of redox-signaling reactive oxygen and nitrogen species such as superoxide and nitric oxide. We generated transgenic plants of Arabidopsis in which XDH was knocked out by introduction of hairpin RNA-expression vector. Expression analysis by reverse transcription-PCR and in-gel staining of XDH activity revealed that transgenic lines efficiently suppressedXDH expression at the transcriptional level, demonstrating that RNA interference was successfully induced. XDH-suppressed transgenic lines exhibitedincreased biomass production during the growth of seedlings.

• Journal of Plant Biotechnology
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• 제37권2호
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• pp.174-185
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• 2010

Full-length cDNAs are essential for the correct annotation of genomic sequences and for the functional analysis of genes and their products. To elucidate the functions of a large population of Chinese cabbage (Brassica rapa) genes and to search efficiently for agriculturally useful genes, we have been taking advantage of the full-length cDNA Over-eXpresser (FOX) gene hunting system. With oligo dT column it purify the each mRNA from the flower organs, leaf and stem tissue. And about 120,000 cDNAs from the library were transformed into $\lambda$-pFLCIII-F vector. Of which 115,000 cDNAs from the library were transformed into T-DNA binary vector, pBigs for transformation study. We used normalized full-length cDNA and introduced each cDNA into Arabidopsis by in planta transformation. Full-length Chinese cabbage cDNAs were expressed independently under the CaMV 35S promoter in Arabidopsis. Selfed seeds were harvested from transgenic Arabidopsis. We had selected 2,500 transgenic plants by hygromycin antibiotic tolerant test, and obtained a number of transgenic mutants. Each transgenic Arabidopsis was investigated in morphological changes, fertility and leaf colour. As a result, 285 possible morphological mutants were identified. Introduced cDNA was isolated by PCR amplification of the genomic DNA from the transgenic mutants. Sequencing result and BLAST analysis showed that most of the introduced cDNA were complete cDNAs and functional genes. Also, we examined the effect of Bromelain on enhancing resistance to soft rot in transgenic Chinese cabbage 'Osome'. The bromelain gene identified from FOX hunting system was transformed into Chinese cabbage using Agrobacterium methods. Transformants were screened by PCR, then RT-PCR and real time PCR were performed to analyze gene expression of cysteine protease in the T1 and T2 generations. The anti-bacterial activity of bromelain was tested in Chinese cabbages infected with soft rot bacteria. The results showed that the over-expressed bromelain gene from pineapple conferred enhanced resistance to soft rot in Chinese cabbage.

• BMB Reports
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• 제43권1호
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• pp.34-39
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• 2010

Expression patterns of OsAREB1 revealed that expression of OsAREB1 gene can be induced by ABA, PEG and heat. Yeast one-hybrid assay demonstrated it can bind to ABA-responsive element (ABRE), which was found in most stress-induced genes. Transgenic Arabidopsis over-expressing OsAREB1 had different responses to ABA and glucose compared to wild-type plants, which suggest OsAREB1 might have a crucial role in these two signaling pathways. Further analysis indicate that OsAREB1 have multiple functions in Arabidopsis. First, OsAREB1 transgenic plants had higher resistance to drought and heat, and OsAREB1 up-regulated the ABA/stress related gene such as RD29A and RD29B. Second, it delayed plant flowering time by down-regulating the expression of flowering-related genes, such as FT, SOC1, LFY and AP1. Due to the dates, OsAREB1 may function as a positive regulator in drought/heat stresses response, but a negative regulator in flowering time in Arabidopsis.