中文摘要：本发明涉及分子生物学及生物检测技术领域，具体公开了基于环介导等温扩增消光技术的转基因玉米T25检测方法。本发明针对转基因玉米T25的pat基因的6个区域设计4条特异性引物，其上下游外引物序列如SEQ?ID?NO.1?2所示，其上下游内引物序列如SEQ?ID?NO.3?4所示。利用环介导等温扩增消光技术使用所述引物对样品的DNA进行扩增，通过反应颜色或吸光值的变化判断是否有扩增产物，从而检测转基因玉米T25。本发明方法具有灵敏度高、特异性强、操作简单、可肉眼判定结果等优点，为转基因玉米T25检测提供了有效、快速的检测方法。
 
外文摘要：NOVELTY - A primer chosen from upstream outer primer comprising a 37 base pair sequence (SEQ ID NO: 1) fully defined in the specification, downstream outer primer comprising a 36 base pair sequence (SEQ ID NO: 2) fully defined in the specification, upstream inner primer comprising a 58 base pair sequence (SEQ ID NO: 3) fully defined in the specification and downstream inner primer comprising a 59 base pair sequence (SEQ ID NO: 4) fully defined in the specification, is new.
USE - The primer is useful for detecting transgenic corn T25 based on a loop-mediated isothermal amplification technique (claimed).
ADVANTAGE - The primer detects transgenic corn T25 simply and rapidly with high sensitivity, strong specificity.
DETAILED DESCRIPTION - INDEPENDENT CLAIMS are included for the following:
(1) method for detecting transgenic corn T25 based on a loop-mediated isothermal amplification technique, involves amplifying DNA extracted from the sample using the above-mentioned primer, and detecting transgenic corn T25 by determining judged presence or absence of amplified product by change of the reaction color or the absorbance value; and
(2) kit comprising the primer.
TF   TECHNOLOGY FOCUS - BIOTECHNOLOGY - Preferred Method: The method involves extracting genome template of the sample to be tested, performing color reaction by mixing the primer with hemin working solution, adding 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) coloring liquid and hydrogen peroxide to obtain a colored reaction mixture, recording color immediately recorded or measuring absorbance at 419 nm, performing loop-mediated isothermal amplification at 60-65 degrees C for 40-60 minutes using the obtain color reaction and genome template, and detecting product, where the hemin working solution comprises 10 mM sodium dihydrogen phosphate, 100 mM potassium chloride, 2 mM magnesium chloride and 0.003% Triton X-100 (RTM: Nonionic surfactant having a hydrophilic polyethylene oxide chain and an aromatic hydrocarbon lipophilic or hydrophobic group).
 
主权项：基于环介导等温扩增消光技术的检测转基因玉米T25的引物，其特征在于，所述引物包括上游外引物、下游外引物、上游内引物和下游内引物，其序列分别如SEQ?ID?NO.1?4所示。
 
申请号：CN201610941424.7
专利号：CN106367524-A
公开/公告号：CN106367524-A
申请/专利权人：中国农业大学
发明/设计人：黄昆仑;许文涛;徐瑗聪;王晨光;罗云波;田文莹
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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