关键词: 转基因作物, 品系特异性检测, 半巢式聚合酶链式反应, 反向聚合酶链式反应, 二重聚合酶链式反应

Abstract: We report the cloning of two flanking sequence of the integrated gene construct of genetically modified maize 59122 by inverse PCR method and the design of even-specific primers based on the left flanking sequence with the aim of developing of a duplex PCR assay for the event-specific transgenic detection of genetically modified maize 59122 using semi-nested PCR, result ing in an amplification fragment of 100 bp in length stretching from the terminator of the pat gene to the 59122 flanking genes. This assay has been successfully applied to detect genetically modified maizes 59122, MON863, MON810, GA21, NK603, genetically modified Roundup Ready soybeans and genetically modified oilseed rape GT73, with high specificity. The optimum concentration of linear DNA in a connection system for detecting genetically modified maize 59122 by this assay was around 1 ng/μL, which exhibited a limit of detection of 0.1% and a sensitivity of 38 copies of haploid genome. Therefore, the developed PCR assay is applicable to detect genetically modified maize and its derivates accurately, fast and efficiently, and can serve to verify routine PCR qualitative detection.

Key words: genetically modified crop, event-specific transgenic detection, semi-nested polymerase chain reaction, inverse polymerase chain reaction, duplex polymerase chain reaction