CAJ | 학술논문

[目的]建立MON89788大豆转化体特异性定性PCR检测方法.[方法]利用TAIL-PCR技术分离MON89788大豆的3＇端旁侧序列,据此序列设计3条特异性引物用于旁侧序列的扩增.经过3轮PCR扩增,获得特异性扩增产物,将此产物回收后克隆到pMD-18T载体上测序,所得序列用Vector NTI分析,并提交NCBI进行序列比对.并对该方法的特异性和灵敏度进行测试.[结果]获得了1 142 bp的3＇端旁侧序列.经Blast检索比对,该序列包括2部分,1～618 bp为载体序列(E9终止子部分序列和LB序列),619～1 142 bp为大豆基因组序列;依据该序列建立的PCR检测方法能特异性地从MON89788大豆中扩增出170 bp的产物,检测灵敏度达到0.05%,约为40个起始模板拷贝.[结论]该研究建立的方法特异性强、灵敏度高,可适用于MON89788大豆检测.
[목적]건립MON89788대두전화체특이성정성PCR검측방법.[방법]이용TAIL-PCR기술분리MON89788대두적3＇단방측서렬,거차서렬설계3조특이성인물용우방측서렬적확증.경과3륜PCR확증,획득특이성확증산물,장차산물회수후극륭도pMD-18T재체상측서,소득서렬용Vector NTI분석,병제교NCBI진행서렬비대.병대해방법적특이성화령민도진행측시.[결과]획득료1 142 bp적3＇단방측서렬.경Blast검색비대,해서렬포괄2부분,1～618 bp위재체서렬(E9종지자부분서렬화LB서렬),619～1 142 bp위대두기인조서렬;의거해서렬건립적PCR검측방법능특이성지종MON89788대두중확증출170 bp적산물,검측령민도체도0.05%,약위40개기시모판고패.[결론]해연구건립적방법특이성강、령민도고,가괄용우MON89788대두검측.
[Objective] The aim was to establish an event-specific qualitative PCR method for transgenic soybean MON89788.[Method] Firstly,the 3'-junction sequence between host plant DNA and integrated DNA of transgenic MON89788 soybean was isolated using thermal asymmetric interlaced-PCR (TAIL-PCR),and the specific PCR primers were designed based on the 3'-junction sequence.Secondly,the specificity and sensitivity of the qualitative PCR detection methods employing these primers were tested.[Result] 1 142-bp 3'-junction sequence was obtained.According to the sequence,event-specific qualitative PCR method was established,amplifying a 170-bp product specifically from MON89788 event,and the limit of detection was 0.05%,approximately 40 initial template copies.[Conclusion] The method was highly specific,sensitive,and suitable for detection of MON89788 event.