中文摘要：本发明涉及分子遗传育种领域，具体提供了一种快速检测LOX转基因小麦的方法及应用该方法的试剂盒，以特异、敏感的外引物F3和B3、内引物FIP和BIP为扩增靶序列。检测方法通过DNA提取和LAMP检测，反应液颜色由紫蓝色变为天蓝色，则为转基因小麦品系。本发明对DNA模板质量要求不高，LAMP检测周期短，检测成本低，操作过程简便，结果分析简单。
外文摘要：NOVELTY - Method for performing loop-mediated isothermal amplification (LAMP) method for detecting LOX transgenic wheat, involves using primers comprising outer forward primer F3 comprising base pair sequence of SEQ ID NO: 1, outer reverse primer R3 comprising base pair sequence of SEQ ID NO: 2, inner forward primer FIP comprising a 43 base pair sequence (SEQ ID NO: 3) fully defined in the specification and inner reverse primer RIP comprising a 39 base pair sequence (SEQ ID NO: 4) fully defined in the specification.
USE - The method or kit is useful for performing loop-mediated isothermal amplification for detecting LOX transgenic wheat (claimed).
ADVANTAGE - The method detects LOX transgenic wheat simply and rapidly with improved specificity and sensitivity, does not require DNA template to be with high quality, and is performed simply and cost-effectively.
DETAILED DESCRIPTION - Method for performing loop-mediated isothermal amplification (LAMP) method for detecting LOX transgenic wheat, involves using primers comprising outer forward primer F3 comprising base pair sequence of SEQ ID NO: 1, outer reverse primer R3 comprising base pair sequence of SEQ ID NO: 2, inner forward primer FIP comprising a 43 base pair sequence (SEQ ID NO: 3) fully defined in the specification and inner reverse primer RIP comprising a 39 base pair sequence (SEQ ID NO: 4) fully defined in the specification. gttcagccagtgctgacaa (SEQ ID NO: 1), and agcctagccaatcttcacaatc (SEQ ID NO: 2). An INDEPENDENT CLAIM is included for kit, which comprises 2.5 mu l 10x LAMP reaction buffer, 1 mu l 8 U/ mu l Bacillus stearothermophilus (Bst) DNA polymerase, 0.5 mu l 10 mu mol/l primer F3, 0.5 mu l 10 mu mol/l primer B3, 1.6 mu l 20 mu mol/l primer BIP, 1.6 mu l 20 mu mol/l primer FIP, 1 mu l DNA template, 4 mu l 5 mol/l betaine, 3.5 mu l 10 mmol/l deoxynucleoside triphosphate, 1 mu l 100 mmol/l magnesium sulfate solution, 1 mu l hydroxynaphthol blue, double-distilled wa! ter (to make up to 25 mu l), and positive control.
主权项：一种快速检测LOX转基因小麦的LAMP方法，其特征在于：其中所采用的引物包括：外引物F3：5'?GTTCAGCCAGTGCTGACAA?3'，其核苷酸序列如SEQ?ID?No.1所示；外引物B3：5'?AGCCTAGCCAATCTTCACAATC?3'，其核苷酸序列如SEQ?ID?No.2所示；内引物FIP：5'?CGCCTGCAGGTCGACCATATGGCTGATGCCATCTAG?ACTATGC?3'，其核苷酸序列如SEQ?ID?No.3所示；内引物BIP：5'?GCCGCAAAGAGGACTAACCGCCACAACACCGAGC?ACCAC?3'，其核苷酸序列如SEQ?ID?No.4所示。
申请号：CN201611090104.1
专利号：CN106520981-A
公开/公告号：CN106520981A
申请日：2016-11-29
公开/公告日：2017-03-22
申请/专利权人：山东农业大学
发明/设计人：孔令让 ; 王彪 ; 杜旭烨
分类号：C12Q1/68
主分类号：C12Q1/68
法律状态：实质审查
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