CAJ | 학술논문

根据7种转基因玉米(Zea mays)Bt11、Bt176、Mon810、Mon863、TC1507、GA21和NK603的重组DNA结构分别设计转化体特异性引物,并对引物特异性及灵敏度进行验证.结果表明,所设计引物特异性强,灵敏度达0.1%,在此基础上以玉米内参照基因(ZSSIIb)作对照,对Bt11、Mon810、TC1507和GA21、NK603、Bt176分别作了2对四重PCR反应.根据插入外源基因与插入位点植物基因组DNA的特异结构,分别设计和筛选了7种转基因玉米转化体特异性Oligo探针,制备转基因玉米的寡核苷酸芯片.结果表明,所设计探针特异性强,灵敏度达0.01%.利用基因芯片检测方法结合多重PCR,建立并优化了1套三重PCR反应体系和两套四重PCR反应体系,可在1张芯片上同时有效地检测及鉴定多种转基因玉米,大大提高了检测的准确率和效率.
근거7충전기인옥미(Zea mays)Bt11、Bt176、Mon810、Mon863、TC1507、GA21화NK603적중조DNA결구분별설계전화체특이성인물,병대인물특이성급령민도진행험증.결과표명,소설계인물특이성강,령민도체0.1%,재차기출상이옥미내삼조기인(ZSSIIb)작대조,대Bt11、Mon810、TC1507화GA21、NK603、Bt176분별작료2대사중PCR반응.근거삽입외원기인여삽입위점식물기인조DNA적특이결구,분별설계화사선료7충전기인옥미전화체특이성Oligo탐침,제비전기인옥미적과핵감산심편.결과표명,소설계탐침특이성강,령민도체0.01%.이용기인심편검측방법결합다중PCR,건립병우화료1투삼중PCR반응체계화량투사중PCR반응체계,가재1장심편상동시유효지검측급감정다충전기인옥미,대대제고료검측적준학솔화효솔.
Based on integrated DNAs structure of seven genetically modified maize(Zea mays),Bt11,Bt176,Mon810,Mon863,TC1507,GA21 and NK603,event specific primers were designed,and the PCR system and program were optimized.The results indicated that the sensitivity of PCR detection was 0.1%.In addition,two multiplex PCR reactions were separately developed by specific fragments with different lengths in Bt11,Mon810,TC1507 and GA21,NK603 and Bt176 which could be amplified simultaneously in one PCR reaction.Meanwhile,maize endogenous reference gene ZSSIIb could also be detected.Based on the unique and specific integration junction sequences between the host plant genome DNA and the integrated gene,a 40mer Oligo probes with high specificity have being developed.Then the transgenic detection DNA chips were got.The results indicated that the sensitivity of microarray detection was 0.01%and the event specific oligonucleotide probes could meet the detecting requirements of sensitivity and specificity as well as high efficiency.One set of 3×multiplex PCR system and two sets of 4×multiplex PCR system for detecting and identifying transgenic maize were set up and optimized.Different genes could be detected specifically in one oligonucleotide microarray using multiplex PCR-gene chip which improved the detection efficiency and accuracy.