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Kong-Sik Shin,Myung-Ho Lim,Hee-Jong Woo,Sun-Hyung Lim,Hong-Il Ahn,Jin-Hyoung Lee,Hyun-Suk Cho,Soon-Jong Kweon,Seok Cheol Suh 한국응용생명화학회 2012 Applied Biological Chemistry (Appl Biol Chem) Vol.55 No.3
Transgenic Chinese cabbage 416-3 was developed in Korea by a transformation event involving modified insectresistant gene (cry1Ac1). To monitor unintended release of genetically modified (GM) Chinese cabbage in the future, as well as to meet GM-labeling requirements, the development of a reliable method for detection of GM cabbage is requisite. To develop qualitative and quantitative polymerase chain reaction methods for the insect-resistant GM Chinese cabbage, a cytosolic glutathione reductase (BcgGR1) gene was used as the endogenous reference gene. Primer pairs CGR-1/-2, amplifying the Chinese cabbage endogenous gene, yielded an expected amplicon of 121bp, whereas no amplified product was observed when DNA samples from seven non-cabbage plants were used as templates. The event-specific primer pairs amplifying the junction site between the endogenous genome sequence and the transferred DNA of GM event 416-3, produced amplicons of desired size by qualitative polymerase chain reaction (PCR) assay. An eventspecific quantitative PCR detection method was established using a TaqMan probe and a standard plasmid as a reference molecule,which contained both endogenous and event-specific sequences. For the validation of this method, three different compositions of w/w mixed samples (containing transgenic DNA at 5, 1, and 0.5%of total DNA in the control samples) were quantified. The precision, expressed as standard deviation (SD) and relative standard deviations (RSD), deviated by 0.03–0.26% and 4.75–8.06%, respectively. These results clearly demonstrate that the PCR methods developed herein can be used for event-specific qualitative and quantitative testings of insect-resistant GM Chinese cabbage.
PCR Detection of Disease Resistant Transgenic Rice for the Biosafety Assessment
Kong Sik Shin,Myung Ho Lim,Hee Jong Woo,Jin Hyoung Lee,Hong Il Ahn,Yang Qin,Hyun Suk Cho 한국육종학회 2012 한국육종학회 심포지엄 Vol.2012 No.07
Regulations in the EU, Japan, Korea, etc. require that foods and feeds made of or derived from genetically modified organisms (GMOs) should be approved and labeled according to a threshold. Recently, disease resistant transgenic rice was developed in Korea, which resulted from the transformation events involving choline kinase gene, OsCK1. In order to monitor unintended release of the developed GM rice in the near future, as well as to meet GM-labeling requirements, the development of a reliable method for detection of disease resistant GM rice is requisite. Here, specific primer pairs for the detection of GMO was designed on the basis of a introduced gene and the flanking junction sequences between a plant DNA and a integrated gene construct, and also SPS gene was used as an endogenous reference material. Specificities of all designed primers were tested through qualitative PCRs. Clearly, target specific amplicons could be detected from disease resistant GM rice event. In addition, the limits of detection (LOD) using the event-specific primers were approximately 0.1% for the disease resistant GM rice line. This result indicated that the developed detection method is suitable for the traceability of disease resistant GM rice, because of using the primer specifically corresponded to the junction site between plant genomic DNA and inserted DNA. Keywords: genetically modified organisms, disease resistant GM rice, PCR detection, event-specific primer
Shin, Kong-Sik,Lim, Myung-Ho,Woo, Hee-Jong,Lim, Sun-Hyung,Ahn, Hong-Il,Lee, Jin-Hyoung,Cho, Hyun-Suk,Kweon, Soon-Jong,Suh, Seok-Cheol The Korean Society for Applied Biological Chemisty 2012 Journal of Applied Biological Chemistry (J. Appl. Vol.58 No.3
Transgenic Chinese cabbage 416-3 was developed in Korea by a transformation event involving modified insect-resistant gene (cry1Ac1). To monitor unintended release of genetically modified (GM) Chinese cabbage in the future, as well as to meet GM-labeling requirements, the development of a reliable method for detection of GM cabbage is requisite. To develop qualitative and quantitative polymerase chain reaction methods for the insect-resistant GM Chinese cabbage, a cytosolic glutathione reductase (BcgGR1) gene was used as the endogenous reference gene. Primer pairs CGR-1/-2, amplifying the Chinese cabbage endogenous gene, yielded an expected amplicon of 121 bp, whereas no amplified product was observed when DNA samples from seven non-cabbage plants were used as templates. The event-specific primer pairs amplifying the junction site between the endogenous genome sequence and the transferred DNA of GM event 416-3, produced amplicons of desired size by qualitative polymerase chain reaction (PCR) assay. An event-specific quantitative PCR detection method was established using a TaqMan probe and a standard plasmid as a reference molecule, which contained both endogenous and event-specific sequences. For the validation of this method, three different compositions of w/w mixed samples (containing transgenic DNA at 5, 1, and 0.5% of total DNA in the control samples) were quantified. The precision, expressed as standard deviation (SD) and relative standard deviations (RSD), deviated by 0.03-0.26% and 4.75-8.06%, respectively. These results clearly demonstrate that the PCR methods developed herein can be used for event-specific qualitative and quantitative testings of insect-resistant GM Chinese cabbage.
연구보문 : 자연과학; 유전자변형 병저항성(OsCK1) 벼에 대한 PCR 검정
신공식 ( Kong Sik Shin† ),이진형 ( Jin Hyoung Lee ),임명호 ( Myung Ho Lim ),우희종 ( Hee Jong Woo ),친양 ( Yang Qin ),조현석 ( Hyun Suk Cho ) 한국국제농업개발학회 2013 韓國國際農業開發學會誌 Vol.25 No.3
Genetically modified (GM) crops generated by agricultural biotechnology are rapidly increasing and cultivated in many countries since they have delivered substantially agronomic, environmental, economic, and social benefits to farmers. The disease resistant transgenic rice, which expresses choline kinase gene (OsCK1), was developed. With the potential problems of safety, the detection method is required as an essential element for the GMO labeling system or GMO traceability of transgenic crops. In this study, gene, construct and event-specific primer pairs with 134, 306 and 243 bp amplicon, respectively, were used for PCR amplification of disease resistant (OsCK1) GM rice and no amplified product was observed from any other crops as templates. The limits of detection (LOD) of qualitative duplex PCR were 0.05% for OsCK1 GM rice using the event-specific primer pair CKRB32-1/02-2. For quantitative detection using TaqMan real-time PCR system, plasmid pSPSCKR as reference molecule was constructed, which contains rice endogenous gene sequence and 5`-junction DNA sequence of OsCK1 GM rice. The absolute detection limit of quantitative PCR method was around 10 copies for one plasmid molecule pSPSCKR. Thereafter, five mixed transgenic rice containing 0.5, 1, 3, 5 and 10% were quantified to evaluate the accuracy and precision of the established real-time PCR system. The bias and the relative deviations were all within the range of 20% for OsCK1 GM rice. These results demonstrate that the developed event-specific qualitative and quantitative PCR methods are acceptable for monitoring and traceability of GM rice.
국내산 복분자 열매에 대한 화학적 조성 및 생리활성 비교
신공식(Kong Sik Shin),박필재(Pill Jae Park),부희옥(Hee Ock Boo),고정연(Jung Youn KO),한성수(Seong Soo Han) 한국자원식물학회 2003 한국자원식물학회지 Vol.16 No.2
복분자 열매의 생리활성 효과를 알아보기 위하여 재배복분자와 야생복분자를 이용하여 실험을 수행하였다. 총페놀성 화합물 함량은 재배복분자 완숙과와 야생복분자 미숙과에서 각각 222, 190mg/g 이었고, 다당체 함량은 야생 북분자 미숙과가 320 unit로 가장 높은 함량을 나타내었다. 복분자 열매의 전자공여능은 야생복분자 미숙과가 100 ㎍/mL 농도에서 95% 이상으로 높은 활성을 보여 미숙과가 완숙과에 비해 높은 전자공여능을 갖는 것으로 나타났다. SOD 유사활성은 야생복분자 미숙과가 81%, 재배복분자 완숙과가 77%를 나타내어, 총페놀성 화합물과 같은 추세를 보였으며, 과산화지질 형성억제능은 모든 처리구에서 대조구인 α-tocopherol과 같은 억제수준을 나타내었다 복분자 열매의 고혈압 완화효과로써 ACE 활성 저해율은 야생복분자 미숙과와 재배복분자 완숙과가 추출농도 1% 범위에서 98% 이상의 높은 ACE 활성 저해율을 보였다. This study was carried out to investigate biological and antioxidative activities on the fruit of bogbunja (Rubus coreanus Miquel). Total contents of phenolic compounds contents in cultivars ripened fruit and immatured wild-type fruit were 222 and 190mg/g, respectively, Polysaccharide contents of immatured wild-type fruit were the highest value of 320U. For EDA analysis, immatured wild-type fruit showed over 95% in 100㎍/mL of sample concentration, which is the the most effective. Levels of SOD-like activities in immatured and cultivar ripened fruits were 81% and 77%, respectively, For the inhibitory effect on lipid peroxidation, all of bogbunja prepared were similar with those of α-tocopherol as control. The inhibition of ACE activities on the water extracts of bogbunja fruit showed over 98%, especially, in immature wild-type and cultivar bogbunja.