中文摘要：本申请公开了一种转基因玉米品系VCO?01981?5检测方法和试剂。本申请的检测方法，包括在PCR反应体系中加矿物油，混匀转移到微滴发生器产生微滴，然后进行PCR反应，结束后，读取每个微滴扩增信号，用QuantaSoft软件计算待测样品的靶标DNA分子数；PCR反应体系中包括品系VCO?01981?5和HMG基因特异引物和探针，两条探针分别采用FAM和HEX荧光标记。本申请的定量检测方法，设计配套使用的内源基因HMG和品系特异性引物探针，采用双通道法，分别定量出两者在玉米基因组中的分子数，计算出转基因玉米品系VCO?01981?5绝对含量，对转基因成分的研究和我国口岸转基因检测具有重要意义。
外文摘要：NOVELTY - Detecting transgenetic corn strain VCO-01981-5 comprises adding oil in the polymerase chain reaction system, mixing uniformly, transferring to the droplet generator, automatically generating a droplet, transferring all the droplets in the reaction tube, carrying out polymerase chain reaction, reading each droplet amplification signal, and using QuantaSoftV1.3.2 software, calculating the sample number of target DNA molecules, the polymerase chain reaction system duplicating real-time fluorescence polymerase chain reaction amplification system.
USE - The method is useful for detecting transgenetic corn strain VCO-01981-5 (claimed).
DETAILED DESCRIPTION - Detecting transgenetic corn strain VCO-01981-5 comprises adding oil in the polymerase chain reaction system, mixing uniformly, transferring to the droplet generator, automatically generating a droplet, transferring all the droplets in the reaction tube, carrying out polymerase chain reaction, reading each droplet amplification signal, and using QuantaSoftV1.3.2 software, calculating the sample number of target DNA molecules, the polymerase chain reaction system duplicating real-time fluorescence polymerase chain reaction amplification system includes specific primer polymerase chain reaction liquid, strain VCO-01981-5, strain VCO-01981-5 specific probe, HMG gene specific primer, HMG gene specific probes and the sample DNA, the forward primer of the VCO-01981-5 specific primer group includes fully defined 5'-aaggccttcagtctactcctcggg-3' (SEQ ID No.1), the reverse primer of the VCO-01981-5 specific primer set includes fully defined 5'-ctagcggccgctactcgagggattt-3' (SEQ ID No.2), the strain VCO-01981-5 specific probe Included fully defined 5'-cgtcaccaagaagatcagtactcaaacac-3' (SEQ ID No.3), the HMG gene specific primer set of a forward primer includes fully defined 5' - ttggactagaaatctcgtgctga-3 ' (SEQ ID No.4), the reverse primer of the HMG gene specific primer set includes as 5'-ctagc! ggccgcta ctcgagggattt-3',(SEQ ID No.5), and strain VCO-01981-5 specific probe includes 5-caatccacacaaacgcacgcgta-3, (SEQ ID No 6), where the 5 end of SEQ ID No.3 is labeled with FAM fluorescent group, the 3 end has BHQ-1 quenching group, the HMG gene specific probe sequence showed in SEQ ID No.6, 5' end with HEX fluorescent group, the 3' end has BHQ-1quenching group. An INDEPENDENT CLAIM is also included for the reagent comprising first primer probe and second primer probe complex, the first primer probe complex for strain VCO-01981-5 specific detection, the second primer probe complex for the HMG gene specific detection, in the first primer probe combination, the forward primer sequence is as shown in SEQ ID No.1, the reverse primer is shown in SEQ ID No.2, the probe sequence is shown SEQ ID No.3, the probe 5' end is marked with FAM fluorescent group, the 3' end has BHQ-1 quenching group, the second primer probe complex, the forward primer is SEQ ID No.4, the reverse primer sequence is as shown in SEQ ID No.5, the probe sequence is as shown in SEQ ID No.6, the probe 5' end has a HEX fluorescent group and the 3' end has BHQ-1 quenching group.
主权项：一种转基因玉米品系VCO?01981?5的检测方法，其特征在于：包括在PCR反应体系中加入矿物油，混匀后，将其转移到微滴发生器上，自动产生微滴，然后将微滴全部转移到反应管中进行PCR反应，PCR反应结束后，读取每个微滴的扩增信号，并采用QuantaSoft?V1.3.2软件计算待测样品的靶标DNA分子数；所述PCR反应体系为双重实时荧光PCR扩增体系，包括PCR反应液、品系VCO?01981?5特异引物组、品系VCO?01981?5特异探针、HMG基因特异引物组、HMG基因特异探针和待测样品的DNA；所述品系VCO?01981?5特异引物组的正向引物为Seq?ID?No.1所示序列，品系VCO?01981?5特异引物组的反向引物为Seq?ID?No.2所示序列，所述品系VCO?01981?5特异探针为Seq?ID?No.3所示序列，所述HMG基因特异引物组的正向引物为Seq?ID?No.4所示序列，HMG基因特异引物组的反向引物为Seq?ID?No.5所示序列，所述HMG基因特异探
申请号：CN201610933272.6
专利号：CN106636340-A
公开/公告号：CN106636340-A
申请日：2016-10-25
公开/公告日：2017-05-10
申请/专利权人：深圳出入境检验检疫局动植物检验检疫技术中心
发明/设计人：凌杏园 ; 潘广 ; 叶奕优 ; 龙海 ; 章桂明 ; 程颖慧 ; 向才玉 ; 谢晓露
分类号：C12Q1/68 ; C12N15/11
主分类号：C12Q1/68
法律状态：实质审查
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