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目的建立针对我国农业部未颁发农业转基因生物安全证书的转基因苜蓿草品系J163品系特异性等温扩增(loop-mediated isothermal amplification,LAMP)检测方法。方法根据转基因苜蓿草品系J163 5’端外源插入片段与苜蓿草基因组DNA之间的邻接区序列设计6条引物,建立了转基因苜蓿草J163品系特异性LAMP检测方法,并对本方法的特异性、灵敏度进行了测定。结果建立的检测方法特异于转基因苜蓿草J163成分检测,检测最低DNA浓度为(1imit of detection,LOD)为16 pg,相当于10拷贝转基因苜蓿草J163基因组DNA。结论本研究建立的转基因苜蓿草J163品系特异性LAMP检测方法特异性好,灵敏度高,能够快速、准确、稳定地对转基因苜蓿草J163成分进行检测分析。
Objective To establish a method for detecting loop-mediated isothermal amplification (LAMP) of transgenic alfalfa line J163 which has not issued the certificate of agricultural genetically modified organism by Ministry of Agriculture. Methods Six primers were designed according to the sequence of the 5 ’exogenous insert of transgenic alfalfa line J163 and the genomic DNA of alfalfa. A specific LAMP assay was established for the transgenic alfalfa J163 line. The specificity, Sensitivity was measured. Results The detection method was specific to the detection of transgenic alfalfa J163. The minimum detectable DNA concentration (LOD) was 16 pg, which was equivalent to 10 copies of genomic DNA of transgenic alfalfa J163. Conclusion The specific LAMP detection method of transgenic alfalfa J163 established in this study has good specificity and high sensitivity, and can detect and analyze the composition of transgenic alfalfa J163 rapidly, accurately and stably.