中文摘要：本发明具体公开了一种快速检测转基因玉米PAT的引物对及试剂盒，所述引物对包括内参引物对和外源基因引物对；所述试剂盒中的引物对包括所述引物对包括内参引物对和外源基因引物对；试剂盒中反应体系组分包括：HRM反应预混液，内参引物对，外源基因引物对，反应模板，去离子水；本发明的组分和配比合理，检测快捷，准确率高，合适用于荧光PCR检测转基因高粱的Zein基因和PAT基因；表明本发明检测的灵敏度更高，达到0.1％的水平，稳定性较高，模板浓度对数值和Cp值之间也呈良好的线性关系R2≥0.96，本发明的检测结果的准确度高达98％左右，由于本发明的简化了操作步骤，缩短检测周期，而且可以实现定量分析，因此具有较高的应用价值。
 
外文摘要：NOVELTY - Primer pair comprises inner primer pair comprising upstream primer ZeinF and downstream primer ZeinR and foreign gene primer pair comprising upstream primer PATF and downstream primer PATR.
USE - The primer pair is useful in kit for detecting transgenic maize PAT (all claimed).
ADVANTAGE - The primer pair enables simple, rapid, cost-effective, stable and highly sensitive and specific detection of PAT, and exhibits accuracy rate of 98% and template concentration between the value and Cp value shows excellent linear relationship of R2 greater than or equal to 0.96.
DETAILED DESCRIPTION - Primer pair comprises inner primer pair and foreign gene primer pair. The inner primer pair comprising upstream primer ZeinF having base pair sequence of actatgccttggtctgtactatgccgag (SEQ ID NO: Not defined) and downstream primer ZeinR having base pair sequence of cgtaagggctgatgattggcataccgtc (SEQ ID NO: Not defined). The foreign gene primer pair comprising upstream primer PATF having base pair sequence of ggaggccaatactgcatggcagaatc (SEQ ID NO: Not defined) and downstream primer PATR having base pair sequence of gctcatcgaaggcctaatggcattcgg (SEQ ID NO: Not defined). INDEPENDENT CLAIMS are included for the following:
(1) kit comprising above-mentioned primer pair and reaction system, where the 25 mu l reaction system comprising 1 times the final concentration of 15 mu l reaction mix HRM, 0.5 pmol/l final concentration of 0.7-1.2 mu l, preferably 1 mu l primer ZeinF, 0.5 pmol/l final concentration of 0.7-1.2 mu l, preferably 1 mu l ZeinR, 0.5 pmol/l final concentration of 0.7-1.2 mu l, preferably 1 mu l PATF, 0.5 pmol/l concentration of 0.7-1.2 mu l, preferably 1 mu l PATR, 1 mu l DNA template and 25 mu l deionized water; and
(2) method of detecting transgenic maize phosphinothricin acetyl transferase (PAT) using kit involves (a) extracting sample DNA, (b) amplifying the extracted sample using PCR and (c) using high-resolution melting curve ana! lyzing t he amplified product and combining with the amplification curve to determine the knot.
 
主权项：一种快速检测转基因玉米PAT的引物对，其特征在于，所述引物对包括内参引物对和外源基因引物对；内参引物对：上游引物ZeinF：ACTATGCCTTGGTCTGTACTATGCCGAG；下游引物ZeinR：CGTAAGGGCTGATGATTGGCATACCGTC；外源基因引物对：上游引物PATF：GGAGGCCAATACTGCATGGCAGAATC；下游引物PATR：GCTCATCGAAGGCCTAATGGCATTCGG。
 
申请号：CN201710064528.9
专利号：CN106755504-A
公开/公告号：CN106755504A
申请/专利权人：深圳汇创联合自动化控制有限公司
发明/设计人：不公告发明人
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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