CAJ | 학술논문

以大豆品种“吉农28”为材料，利用RT－PCR技术获得GmCHR基因序列，并选择其中918 bp的开放阅读框，以pBI121为基础载体，构建了由35S启动子驱动的GmCHR植物表达载体，转入烟草中并对17株抗卡那霉素的转基因烟草进行PCR、GUS染色、Southern杂交检测，对植株中GmCHR基因mRNA表达量进行荧光定量PCR检测，采用高效液相色谱技术测定转基因植株中查尔酮还原酶催化产物异甘草素的含量。结果表明：GmCHR基因成功整合到烟草基因组中并能够成功表达；在转基因植株中目的基因的mRNA均有表达，且不同植株间表达量存在差异；转化烟草叶部组织中异甘草素的平均含量为（7?528±0?018）μmol／g，而转空载体烟草中未检测出异甘草素。
이대두품충“길농28”위재료，이용RT－PCR기술획득GmCHR기인서렬，병선택기중918 bp적개방열독광，이pBI121위기출재체，구건료유35S계동자구동적GmCHR식물표체재체，전입연초중병대17주항잡나매소적전기인연초진행PCR、GUS염색、Southern잡교검측，대식주중GmCHR기인mRNA표체량진행형광정량PCR검측，채용고효액상색보기술측정전기인식주중사이동환원매최화산물이감초소적함량。결과표명：GmCHR기인성공정합도연초기인조중병능구성공표체；재전기인식주중목적기인적mRNA균유표체，차불동식주간표체량존재차이；전화연초협부조직중이감초소적평균함량위（7?528±0?018）μmol／g，이전공재체연초중미검측출이감초소。
GmCHR gene sequence was isolated from soybean variety“jinong 28” by RT-PCR tech?nique. The 918 bp open reading frame was selected in that sequence. On the basis of pBI121 carri?er, a GmCHR recombinant botany expression vector driven by 35S promoter was constructed and transferred into tobacco. Seventeen plants of kan resistant transgenic tobacco were identified by mo?lecular biology method of PCR and Southern blotting. Tested positive transgenic tobacco was per?formed with GUS activity assay. Expression value of the gene in different plants was determined by real?time fluorescent quantitative PCR and the content of isoliquiritigenin catalyticed product by the enzyme was identified by HPLC technology. The results indicated that GmCHR gene had been inte?grated into the tobacco genome and expressed successfully. The expression value of the gene in the leaves of different transgenic plants was significantly different. Average content of isoliquiritigenin catalyzed by enzyme in tobacco leaf tissue was (7?528±0?018)μmol/g, but isoliquiritigenin was not detected in empty vehicle transfected tobacco.