[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...[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.展开更多
[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant D...[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant DNA integrated DNA of MON89788 soybean were amplified independently,and the three fragments were cloned into the cloning vector pMD18-T in order through molecular manipulation method to construct pMD-LM3M5,the applicability of the constructed novel PRM was tested. [Result] Sequencing confirmation result showed that the PRM was 3 700 bp in length,containing 1 029 bp of recombined DNA fragment. The limits of qualitative detection of the PRM were 10 copies. [Conclusion] The PRM constructed in this study was suitable for the identification of MON89788 event.展开更多
[目的]为完善我国转基因检测方法体系,建立转基因大豆MON87751品系特异性实时荧光聚合酶链式反应(real time polymerase chain reaction,PCR)检测方法。[方法]根据MON87751的3′端邻接区序列设计特异性引物和探针,建立MON87751品系特异...[目的]为完善我国转基因检测方法体系,建立转基因大豆MON87751品系特异性实时荧光聚合酶链式反应(real time polymerase chain reaction,PCR)检测方法。[方法]根据MON87751的3′端邻接区序列设计特异性引物和探针,建立MON87751品系特异性实时荧光PCR检测方法,并测定该方法的灵敏度、特异性及可重复性。[结果]灵敏度测试显示,其定量下限为40拷贝;重复性试验显示其相对标准偏差在可接受范围内。[结论]该研究建立的MON87751品系特异性实时荧光PCR检测方法特异性良好,灵敏度高,有良好的可重复性,适合对转基因大豆MON87751品系进行检测鉴定。展开更多
The main advantage of digital PCR(dPCR) is that it facilitates absolute quantification of the target without reference to the standard/calibration curve.Crystal droplet dPCR has a three-color staining detection functi...The main advantage of digital PCR(dPCR) is that it facilitates absolute quantification of the target without reference to the standard/calibration curve.Crystal droplet dPCR has a three-color staining detection function,which enables multiplex PCR reaction.In this study,this technique was used to establish triple dPCR detection for the genetically modified soybean MON87708 × MON89788 with stacked traits.Specific absolute quantitative detection was accomplished for the genomic DNA extracted from the homogenized seeds of GM stack MON87708× MON89788 soybean.Our results can serve as a reference for the absolute quantitative detection of stacked events of genetically modified crops.展开更多
基金Supported by Important National Science & Technology Specific Pro-jects(2008ZX08012-001)~~
文摘[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.
基金Supported by Major Projects of Cultivating New Varieties by Trans-genic Technology (2008ZX08012-001)~~
文摘[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant DNA integrated DNA of MON89788 soybean were amplified independently,and the three fragments were cloned into the cloning vector pMD18-T in order through molecular manipulation method to construct pMD-LM3M5,the applicability of the constructed novel PRM was tested. [Result] Sequencing confirmation result showed that the PRM was 3 700 bp in length,containing 1 029 bp of recombined DNA fragment. The limits of qualitative detection of the PRM were 10 copies. [Conclusion] The PRM constructed in this study was suitable for the identification of MON89788 event.
文摘[目的]为完善我国转基因检测方法体系,建立转基因大豆MON87751品系特异性实时荧光聚合酶链式反应(real time polymerase chain reaction,PCR)检测方法。[方法]根据MON87751的3′端邻接区序列设计特异性引物和探针,建立MON87751品系特异性实时荧光PCR检测方法,并测定该方法的灵敏度、特异性及可重复性。[结果]灵敏度测试显示,其定量下限为40拷贝;重复性试验显示其相对标准偏差在可接受范围内。[结论]该研究建立的MON87751品系特异性实时荧光PCR检测方法特异性良好,灵敏度高,有良好的可重复性,适合对转基因大豆MON87751品系进行检测鉴定。
文摘The main advantage of digital PCR(dPCR) is that it facilitates absolute quantification of the target without reference to the standard/calibration curve.Crystal droplet dPCR has a three-color staining detection function,which enables multiplex PCR reaction.In this study,this technique was used to establish triple dPCR detection for the genetically modified soybean MON87708 × MON89788 with stacked traits.Specific absolute quantitative detection was accomplished for the genomic DNA extracted from the homogenized seeds of GM stack MON87708× MON89788 soybean.Our results can serve as a reference for the absolute quantitative detection of stacked events of genetically modified crops.
