온주밀감(Citrus unshiu Marc. var. Okitsu) 미분화 종바로부터 callus 유기 및 증식

진성범,홍경애,부경환,이도승,류기중 제주대학교 방사능이용연구소 2000 연구보고 Vol.14 No.-

조직배양을 통한 기관분화나 식물체 재분화 가 어려운 것으로 알려져 있는 온주밀감(Citrus unshiu Marc. var.Okitsu) 미분화 종자로부터 callus 유기 조건을 확립하였다. 온주밀감 성숙과실의 미분화 종자를 MT(Murashige and Tucker, 1969) 기본배지에 치상 6주 후에 미숙종자 양쪽 가장자리에 유기된 조그만white callus를 gibberellin 1mg/L, adenine 25mg/L, malt extract 500mg/L가 첨가된 MT 기본배지에 치상 하여 2주간격으로 계대배양하여 캘러스를 유지할 수 있었고, EME(Grosser and Gmitter, 1990a ) 배지에 4주 간격으로 계대배양함으로써 증식시킬 수 있었다.

CLONING AND EXPRESSION OF E.COLI ORNITHINE TRANSCARBAMYLASE GENE, argl

Riu,Key-Zung,Koh,Young-Hwan,Kim,Chan-Sik,Lee,Sun-Joo 제주대학교 방사능이용연구소 1994 연구보고 Vol.8 No.-

E. coli ornithine transcarbamylase is the enzyme which catalyzes the citrullin biosynthesis from ornithine and carbamyl phosphate. To facilitate the purification of enzyme which will be used for many purposes, cloning and expression of E. coli argI gene, ornithine transcarbamylase gene, have been conducted. argI has been amplified from genomic DNA of E. coli DH5a strain by PCR method, and cloned in prokaryotic expression vector, pKK223-3. The enzyme expressed has been purified by salt fractionation, heat denaturation and affinity chromatography. The result of SDS PAGE shows a single band of the purified enzyme. Kinetic data for expressed enzyme give almost same results as those of the wild type enzyme. Vmax of the enzyme 1.0x105 min-¹, and Kms of ornithine and carbamyl phosphate are 0.35 mM and 0.06 mM, respectively. On the other hand, TB2 cell has been obtained by plasmid curing. This strain does not have argI gene and resulting ornithine transcarbamylase activity. The strain has been prepared for site-directed mutagenesis work.

Construction of Citrus Transgenic Plant with Fatty Aicd Desaturase Gene

Riu, Key Zung,Jin, Seong Beom,Boo, Kyung Hwan,Lee, Do Seung,Chae, Hyun Byung,Song, Seong Jun 한국농화학회 1999 Applied Biological Chemistry (Appl Biol Chem) Vol.42 No.3

The transgenic plant of Citrus species (Citrus aurantium L.) was constructed with a fatty acid desaturase gene using microprojectile bombardment transformation system. The DNA of a fatty acid desaturase gene, fad7, constructed in pBI121 was coated onto tungsten particles (1.1 ㎛) and introduced into callus cells by bombarding with 1100 psi of helium pressure, l/4 in of gap distance, 7.0 ㎝ of target distance and 27 in Hg of chamber vacuum. The bombarded cells were selected on the medium containing kanamycin. The selected cells were successfully regenerated into plantlets via somatic embryogenesis on the media containing plant growth regulators. The results of polymerase chain reaction analysis of genomic DNAs from the putative transformants showed that the introduced DNAs of fad7 were present in both the selected callus cells and the regenerated plantlets.

Anti-bacterial and Anti-viral Activities of Extracts from Terminalia chebula Barks

Key Zung Riu,Doseung Lee,부경환,Jin-Kyu Woo,Fangmeng Duan,이근화,권택규,이효연,이동선 한국응용생명화학회 2011 Applied Biological Chemistry (Appl Biol Chem) Vol.54 No.2

Effects of methanol extract of Terminalia chebula on antibacterial activity and the generation of superoxide radical in Bacillus subtilis, as well as on syncytium formation and cytopathic activity in virus-infected baby hamster kidney cells were examined. The extract effectively inhibited syncytium formation in a concentration-dependent manner, and infectious virus production was markedly reduced. However, glycoprotein synthesis was not affected. These results collectively indicate that methanol extract of T. chebula potentially inhibit glycosylation by acting as a suppressor of intracellular glycosylation trafficking.

Escherichia coli 오르니틴 트란스카바밀라제의 유전자 argⅠ 의 클로닝 및 발현

류기중(Key Zung Riu),유장걸(Zang Kual U),고영환(Young Hwan Ko),김찬식(Chan Shik Kim),송성준(Sung Jun Song),오영선(Young Seon Oh),이선주(Sun Joo Lee) 한국응용생명화학회 1995 Applied Biological Chemistry (Appl Biol Chem) Vol.38 No.2

Escherichia Coli ornithine transcarbamylase is the enzyme which catalyzes the L-citrulline biosynthesis from L-ornithine and carbamyl phosphate. To facilitate the purification of enzyme which will be used for many biochemical studies such as structure and function relationships and catalytic mechanisms, the cloning and expression of E. coli argI gene for ornithine transcarbamylase was conducted. argI was amplified from genomic DNA of E. coli strain of DH5α, by polymerization chain reaction (PCR) method. The amplified argI gene was ligated to the prokaryotic expression vector pKK223-3 and used for transformation of E. coli TB2 which was deficient of ornithine transcarbamylase. The over-produced enzyme by the tnansformant was purified by ammonium sulfate fractionation, heat denaturation and affinity chromatography. The result of SDS denaturation gel electrophoresis for the purified enzyme showed a single band of about 38 kDa of ornithine transcarbamylase. Kinetic data for the expressed enzyme gave almost the scone values as those of the wild type enzyme. The k_(cat), of the enzyme was 1.0×10^5 min^(-1) and K_ms for ornithine and carbamyl phosphate were 0.35 mM and 0.06 mM, respectively.

Agrobacterium rhizogenes 를 이용한 Populus tremuloides 의 형질전환

류기중(Key Zung Riu),소인섭(In Sup So),유장걸(Zang Kual U),고영환(Yong Hwan Ko),이선주(Sun Joo Lee),(Wesley P . Hackett) 한국응용생명화학회 1995 Applied Biological Chemistry (Appl Biol Chem) Vol.38 No.2

Several factors affecting on transformation efficiency were studied to establish a Agrobacterium rhizogenes mediated system for the transformation of Populus species, and We could obtaine tansgenic plantlets expressing the introduced gene. Leaf sections were more sensitive than stem sections to kanamycin and thought to be better material for transformant screening. The bacterial density did not affect on the efficiency of transformation over the range of 4×10^5∼7×10^9 cfu. The optimum period for co-cultivation was one day or shorter. Both of the optimum concentrations of cefotaxime and ampicillin in the medium were 250 ㎍/㎖ for elimination of bacteria from the inoculated leaf sections. The addition of acetosyringone in the bacterial culture medium increased transformation rate, and the highest rate was obtained at 50 μM of acetosyringone. The transformed galls could be selectively induced and gown on the growth regulator-free medium or on the medium containing 100 ㎍/㎖ or higher contrition of kanamycin. The roots were induced from the galls incited by A rhizogenes within 3 weeks on the growth regulator-free medium as well as on the medium containing growth regulators. The plantlets were regenerated from the galls cultured for 6 weeks on the medium containing 0.05 ㎎/㎖ of NAA and 0.5 ㎎/㎖ of BA. The expressions of the introduced opine gene in the transformed galls and plantlets were confirmed by the analysis of agropine and mannopine.

식물 원형질체에서의 Marker Gene 삽입

류기중,소인섭,홍경애,유장걸 제주대학교 방사능이용연구소 1994 연구보고 Vol.8 No.-

The neomycin phosphotransferase Ⅱ gene(nptⅡ) was introduced into geranium(Pelargonium zonale hybrids) protoplast by using PEG or electroporation method. The presence of the introduced DNA in the protoplast and the expressions of the gene in the transformed cells were examined. The presence of the nptⅡ DNA in the protoplasts were detected by polymerase chain reaction. The expressions of nptⅡ gene in the transformed cells were confirmed by the NPT-Ⅱassay.

Petroleum Spray Oil 살포가 감귤 잎의 광합성관련 특성에 미치는 영향

박진희,류기중,강시용,김판기 한국환경농학회 2001 한국환경농학회지 Vol.20 No.3

PSO (petroleum spray oil) 살포가 감귤잎에 미치는 광합성 관련 생리적 영향을 밝히고자 4 년생 온주밀감(궁천조생)에 PSO를 살포한 후 광합성, 증산량, 기공전도도 및 엽록소형광 등의 변화에 판하여 검토하였다. 시험은 ①PSO 0.33% 및 1.0% 농도별 처리, ②다른 주야간 온도처리(34/24℃, 30/20℃ 및 28/16℃)하에서의 PSO 처리, 그리고 ③ PSO 처리후 일시적인 고온(50℃, 10시간) 처리 등 3개로 나누어서 실시하였다. 그 결과, PSO를 처리하면 감귤 잎에 유침상이 발생하는데 처리후 30∼40일후에는 무시해도 좋을 정도로 사라졌다. 0.33% PSO 처리시에는 광합성, 증산량 및 염색도 등에 별 영향이 없으며, 1% 처리시에는 기공전도도, 증산율, 광합성속도 등이 저해되다가 살포 20일 후에는 회복되는 경향을 나타냈다. 온도 처리별 PSO 처리의 영향은 고온구 (34/24℃)에 1.0% PSO를 처리했을 경우, 처리 7일 이후에 엽록소 형광(Fv/Fm) 값이 저하하였다. 또한 PSO 처리후 인위적으로 고온(50℃, 10시간)처리를 하였을 경우, 1.0% PSO와 디치를 혼용처리한 나무의 낙엽이 현저하게 증가하였으며, 남은 잎에서도 생리적 형질의 저해가 인정되었다. 이러한 결과 0.33% PSO 처리에서는 감귤 잎의 광합성 관련 특성에 별 영향이 없으나, 1.0% PSO 이상의 살포의 경우 한동안 일부 생리적 기능의 저해를 보였으며, 고온조건이거나 디치와 같은 약제와 혼용할 경우에는 생리적 기능저해가 더욱 심해지는 것으로 나타났다. Recently, petroleum spray oil(PSO) has been used to control key pests in integrated pest management (IPM) of citrus and other orchards in Australia and USA. In order to clarify the influences of a newly developed PSO (D-C Tron Plus^ⓡ) on citrus leaves, 0.33% or 1.0% of PSO were sprayed to potted 4-year-old citrus trees under some kinds of condition, and then the changes of photosynthesis, transpiration, stomatal conductance and chlorophyll fluorescence(Fv/Fm) were determined. When sprayed with 1.0% PSO, the photosynthetic rate, transpiration and stomatal conductance of citrus leaves were decreased by 20%, and then recovered in 20 days after treatment (DAT), while there were little influences by the spray of 0.33% PSO. The value of Fv/Fm decreased more under the 34/24℃ temperature condition than that of under the 30/20℃ and 28/16℃ condition. The high temperature (50℃ for 10 hours)-treated trees sprayed with PSO 1.0% or PSO 1.0% plus dithianon 1/2000 dilution showed not only the increase of rate in dropped leaf but also the reduced photosynthesis and Fv/Fm compared with 30/20℃ temperature-treated ones. From the results of this study, the spray of 1.0% PSO can inhibit the physiological activities in citrus leaf, particularly under high temperature condition after spray or the mixing-spray with a fungicide (dithianon WP, 75%).

대두(Glycine max) protoplast의 세포벽재생에 대한 benzyladenine의 영향

박창규,류기중 濟州大學校 放射能利用硏究所 1992 연구보고 Vol.7 No.-

대두(Glycine max)의 β-1,3-glucanase를 분리동정하고 benzyladenine(BA)이 이효소의 세포내 함량과 활동도에 미치는 영향을 조사하였다. 또 세포벽의 callose함량과 protoplast의 세포벽재생에 미치는 BA의 영향을 조사하여, cytokinin이 식물의 세포벽재생을 촉진하는 기능이 있음을 확인하고 세포벽재생에 있어서 cytokinin의 작용기구를 검토하였다. 대두 β-1,3-Glucanase는 21KD의 polypeptide로 동정되었는데 이 polypeptide의 세포내함량과 효소활성은 BA처리에 의하여 저하되었다. 그리고 callus세포벽의 callose함량과 protoplast의 세포벽 재생율이 BA 처리에 의하여 증가되었다. 이 결과들은 cytokinin이 세포의 β-1,3- glucanase 수준을 저하시켜 callose분해를 억제함으로써 세포벽 재생을 촉진할 수 있음을 보여주었다. A β-1,3-glucanase of soybean(Glycine max) was isolated, and the effects of benzyladenine(BA) on celluar levels of the enzyme content and activity were studied. The effects of BA on callose content in cell wall and wall regeneration of protoplasts were also studied to show promoting effect of cytokinin in cell wall regeneration and to elucidate action mode of cytokinin. The palypeptide of 21KD was identified as β-1,3-glucanase, and the cellular content and activity of this polypeptide were decreased by BA treatment. The callose content in cell wall of callus and the wall regeneration of protoplasts were increased by BA treatment. These results indicate that cytokinin promotes cell wall regeneration by inhibition of callose degradation via decreasing β-1,3-glucanase level in cell.

대두(Glycine max) protoplast 의 세포벽재생에 대한 benzyladenine 의 영향

박창규,류기중 한국농화학회 1992 Applied Biological Chemistry (Appl Biol Chem) Vol.35 No.6

A β-1,3-glucanase of soybean (Glycine max) was isolated, and the effects of benzyladenine(BA) on celluar levels of the enzyme content and activity were studied. The effects of BA on callow content in cell wall and wall regeneration of protoplasts were also studied to show promoting effect of cytokinin in cell wall regeneration and to elucidate action mode of cytokinin. The polypeptide of 21 kD was identified as β-1,3-glucanase, and the cellular content and activity of this polypeptide were decreased by BA treatment. The callose content in cell wall of callus and the wall regeneration of protoplasts were increased by BA treatment. These results indicate that cytokinin promotes cell wall regeneration by inhibition of callose degradation via decreasing β-1,3-glucanase level in cell.