College of life science, Northeast Agricultural University of China, Harbin, People's Republic of China.
PLoS One. 2009 Aug 18;4(8):e6679. doi: 10.1371/journal.pone.0006679.
Transgenic animals have been used for years to study gene function, produce important proteins, and generate models for the study of human diseases. However, inheritance and expression instability of the transgene in transgenic animals is a major limitation. Copy number and promoter methylation are known to regulate gene expression, but no report has systematically examined their effect on transgene expression. In the study, we generated two transgenic pigs by somatic cell nuclear transfer (SCNT) that express green fluorescent protein (GFP) driven by cytomegalovirus (CMV). Absolute quantitative real-time PCR and bisulfite sequencing were performed to determine transgene copy number and promoter methylation level. The correlation of transgene expression with copy number and promoter methylation was analyzed in individual development, fibroblast cells, various tissues, and offspring of the transgenic pigs. Our results demonstrate that transgene expression is associated with copy number and CMV promoter methylation in transgenic pigs.
转基因动物已被用于研究基因功能、生产重要蛋白质和建立人类疾病模型多年。然而，转基因在转基因动物中的遗传和表达不稳定是一个主要的限制。已知拷贝数和启动子甲基化可调节基因表达，但尚无报道系统地研究它们对转基因表达的影响。在本研究中，我们通过体细胞核移植（SCNT）生成了两个表达绿色荧光蛋白（GFP）的转基因猪，该蛋白由巨细胞病毒（CMV）驱动。通过绝对定量实时 PCR 和亚硫酸氢盐测序来确定转基因拷贝数和启动子甲基化水平。在个体发育、成纤维细胞、各种组织和转基因猪的后代中，分析了转基因表达与拷贝数和启动子甲基化的相关性。我们的结果表明，转基因在转基因猪中的表达与拷贝数和 CMV 启动子甲基化有关。
