Department of Life Science, Dongguk University-Seoul, Goyang, Republic of Korea.
Department of Genetics and Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York, United States of America.
PLoS One. 2019 Dec 4;14(12):e0224533. doi: 10.1371/journal.pone.0224533. eCollection 2019.
In transgenic research, selection markers have greatly facilitated the generation of transgenic animals. A prerequisite for a suitable selection marker to be used along with a test gene of interest is that the marker should not affect the phenotype of interest in transformed animals. One of the most common selection markers used in C. elegans transgenic approaches is the rol-6 co-injection marker, which induces a behavioral roller phenotype due to a cuticle defect but is not known to have other side effects. However, we found that the rol-6 co-injection marker can cause expression of GFP in the test sequence in a male-specific interneuron called CP09. We found that the rol-6 gene sequence included in the marker plasmid is responsible for this unwanted expression. Accordingly, the use of the rol-6 co-injection marker is not recommended when researchers intend to examine precise expression or perform functional studies especially targeting male C. elegans neurons. The rol-6 sequence region we identified can be used to drive a specific expression in CP09 neuron for future research.
在转基因研究中，选择标记极大地促进了转基因动物的产生。选择标记与感兴趣的测试基因一起使用的一个前提条件是，标记不应影响转化动物中感兴趣的表型。在秀丽隐杆线虫转基因方法中最常用的选择标记之一是 rol-6 共注射标记，由于角质层缺陷，该标记会诱导行为滚轴表型，但据知它没有其他副作用。然而，我们发现 rol-6 共注射标记可以在称为 CP09 的雄性特定中间神经元中使测试序列中的 GFP 表达。我们发现标记质粒中包含的 rol-6 基因序列负责这种不需要的表达。因此，当研究人员打算检查精确表达或进行专门针对雄性秀丽隐杆线虫神经元的功能研究时，不建议使用 rol-6 共注射标记。我们鉴定的 rol-6 序列区域可用于未来研究中在 CP09 神经元中驱动特定表达。
