In 2017, various orange coloured petunia on the market turned out to be genetically modified (GM) without an official authorization for commercialization. Sequence analysis suggested these undeclared plants most probably originated from a plant transformation experiment performed in the 1980s. For a deeper understanding how GM petunia entered classical breeding programmes worldwide, and whether they originated from a single source or not, we undertook a molecular genetic characterization of the T-DNA integration sites in different GM petunia cultivars and breeding lines. By means of genome walking, we isolated different T-DNA sequences, which are located at the junctions between the T-DNA(s) and the petunia DNA. Based on the results obtained we conclude that there are at least two T-DNA copies of different lengths. This is supported by Southern blot analysis. For T-DNA1, the 3′-junction sequence was isolated, whereas the 5′-junction remained unclear. In contrast, for T-DNA2, the 5′-junction sequence was isolated, whereas the sequence isolated from the 3′-region consists only of T-DNA, but did not include the junction from the T-DNA to the petunia DNA. We developed primers for event-specific PCRs and screened a set of three orange GM petunia cultivars and 126 GM offspring from a commercial breeding program. We show that both T-DNA copies are present in all our tested GM petunia samples, which underpins the assumption of a single transgenic origin of the undeclared GM petunia. Most likely, the two T-DNAs are integrated in close proximity into the petunia genome.

2017年，市场上各种橙色矮牵牛竟然未经官方授权进行商业化转基因（GM）。序列分析表明这些未申报的植物很可能起源于1980年代进行的植物转化实验。为了更深入地了解转基因矮牵牛如何进入全球经典育种计划，以及它们是否起源于单一来源，我们对不同的转基因矮牵牛品种和育种系中的T-DNA整合位点进行了分子遗传学表征。通过基因组步移，我们分离了不同的T-DNA序列，它们位于T-DNA与矮牵牛DNA之间的交界处。根据获得的结果，我们得出结论，至少有两个不同长度的T-DNA拷贝。Southern印迹分析支持了这一点。对于T-DNA1，分离了3'接头序列，而5'接头仍然不清楚。相反，对于T-DNA2，分离了5'-接合序列，而从3'-区域分离的序列仅由T-DNA组成，但不包括从T-DNA至矮牵牛DNA的接合。我们开发了用于事件特异性PCR的引物，并从商业育种计划中筛选了三个橙色的GM矮牵牛品种和126个GM后代。我们表明，所有我们测试的GM矮牵牛样品中都存在两个T-DNA拷贝，这支持了未申报的GM矮牵牛的单一转基因起源的假设。最有可能的是，两个T-DNA紧密整合到矮牵牛基因组中。而5'-结仍然不清楚。相反，对于T-DNA2，分离了5'-接合序列，而从3'-区域分离的序列仅由T-DNA组成，但不包括从T-DNA至矮牵牛DNA的接合。我们开发了用于事件特异性PCR的引物，并从商业育种计划中筛选了三个橙色的GM矮牵牛品种和126个GM后代。我们表明，所有我们测试的GM矮牵牛样品中都存在两个T-DNA拷贝，这支持了未申报的GM矮牵牛的单一转基因起源的假设。最有可能的是，两个T-DNA紧密整合到矮牵牛基因组中。而5'-结仍然不清楚。相反，对于T-DNA2，分离了5'-接合序列，而从3'-区域分离的序列仅由T-DNA组成，但不包括从T-DNA至矮牵牛DNA的接合。我们开发了用于事件特异性PCR的引物，并从商业育种计划中筛选出三个橙色的GM矮牵牛栽培品种和126个GM后代。我们表明，所有我们测试的GM矮牵牛样品中都存在两个T-DNA拷贝，这支持了未申报的GM矮牵牛的单一转基因起源的假设。最有可能的是，两个T-DNA紧密整合到矮牵牛基因组中。但不包括T-DNA与矮牵牛DNA的连接。我们开发了用于事件特异性PCR的引物，并从商业育种计划中筛选出三个橙色的GM矮牵牛栽培品种和126个GM后代。我们显示所有我们测试的GM矮牵牛样品中都存在两个T-DNA拷贝，这支持了未申报的GM矮牵牛的单一转基因起源的假设。最有可能的是，两个T-DNA紧密整合到矮牵牛基因组中。但不包括T-DNA与矮牵牛DNA的连接。我们开发了用于事件特异性PCR的引物，并从商业育种计划中筛选出三个橙色的GM矮牵牛栽培品种和126个GM后代。我们表明，所有我们测试的GM矮牵牛样品中都存在两个T-DNA拷贝，这支持了未申报的GM矮牵牛的单一转基因起源的假设。最有可能的是，两个T-DNA紧密整合到矮牵牛基因组中。