中文摘要：本发明提供一种基于环介导等温扩增消光技术的转基因玉米BT176检测方法，首先针对转基因玉米BT176转化事件3’端基因的部分区域设计一套LAMP引物组合，包括BT176?F3/B3和BT176?FIP/BIP(Seq?ID?No:1?4)，在所述引物组合基础上结合环介导等温扩增消光技术检测转基因玉米BT176转化事件。本方法具有灵敏度高、特异性强、操作简单，结果易于观察等优点，为转基因玉米BT176检测提供了有效、快速的检测方法，非常适于基层检测使用。
外文摘要：NOVELTY - A loop-mediated isothermal amplification (LAMP) primer combination for detecting transgenic maize BT176, is new. The LAMP primer combination comprises outer forward primer BT176-F3 having a 35 base pair sequence (SEQ ID NO: Not defined) fully defined in the specification, and outer reverse primer BT176-B3 having 36 base pair sequence (SEQ ID NO: Not defined) fully defined in the specification based on transformation event 3' end sequence design of transgenic maize BT176.
USE - The LAMP primer combination is useful for detecting transgenic maize BT176 (claimed).
ADVANTAGE - The LAMP primer combination detects transgenic maize BT176 in a simple and rapid manner with high sensitivity and specificity.
DETAILED DESCRIPTION - A loop-mediated isothermal amplification (LAMP) primer combination for detecting transgenic maize BT176, is new. The LAMP primer combination comprises outer forward primer BT176-F3 having a 35 base pair sequence (SEQ ID NO: Not defined) fully defined in the specification, and outer reverse primer BT176-B3 having 36 base pair sequence (SEQ ID NO: Not defined) fully defined in the specification, and inner forward primer BT176-FIP having a 56 base pair sequence (SEQ ID NO: Not defined) fully defined in the specification, and inner reverse primer BT176-BIP having a 56 base pair sequence (SEQ ID NO: Not defined) fully defined in the specification based on transformation event 3' end sequence design of transgenic maize BT176. INDEPENDENT CLAIMS are included for the following:
(1) kit comprising LAMP primer combination; and
(2) method for detecting transgenic maize BT176 based on LAMP, involves grinding sample into powder, and extracting genomic DNA using cetyl trimethylammonium bromide (CTAB), observing color of the reaction mixture and measuring optical density (OD) value at 419 nm using UV spectrophotometer, and performing LAMP using extracted genomic DNA and primer combination and analyzing amplified products.
TF   TECHNOLOGY FOCUS - BIOTECHNOLOGY - Preferred Kit: The kit further comprises dinucleotide triphosphate, Bst DNA polymerase, betaine, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) coloring solution, hemin working solution, hydrogen peroxide solution, and standard positive template.
主权项：用于检测转基因玉米BT176的LAMP引物组合，其特征在于，所述引物组合根据转基因玉米BT176转化事件3’端序列设计，包括：外侧正向引物BT176?F3：5’?CTGGGAGGGAGGGAGGGTTCAAGCACGGGAACTGG?3；’外侧反向引物BT176?B3：5’?CTGGGAGGGAGGGAGGGGGAGAGGGAGAGAGAGGGA?3’；以及内侧正向引物BT176?FIP：5’?GATCTCGGTGACGGGCAGGACTGGGAGGGAGGGAGGGGTTTCTGGCAGCTGGACTT?3；’内侧反向引物BT176?BIP：5’?TCTCCTCCATTGATGCACGCCCTGGGAGGGAGGGAGGGGGGGAAGGGAGAAACGGT?3’。
申请号：CN201610909450.1
专利号：CN106367520-A
公开/公告号：CN106367520-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;陈冰
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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