Kong Q R, Wu M L, Zhang L, Wang F, Yin Z, Mu Y S, Liu Z H
Northeast Agricultural University of China, College of Life Science, Harbin, China.
Cell Mol Biol (Noisy-le-grand). 2011 Jun 23;57 Suppl:OL1505-12.
Transgenic technology has been used for years to study gene function, produce important proteins, and generate models for the study of human diseases. However, the efficiency of producing transgenic animal lines that retain normal function is extremely low. The low efficiency can be mainly attributed to the integrated transgene. A further understanding of the effects of transgene integration on transcription and epigenetic modification of the host genome would improve the transgenic efficiency. Therefore, we utilized three transgenic pigs produced by SCNT expressing GFP, to identify alterations of transcription, DNA methylation and histone acetylation resulting from integration of the GFP gene. Multiple copies of the transgene integrated into a single site of the three transgenic pigs were verified by TAIL—PCR and the integration sites were different in each pig. We observed that the integrated transgene frequently resulted in significantly low transcription of flanking sequences in various tissues of transgenic pigs in comparison with wild—type pigs. Corresponding with the low transcription, DNA hypermethylation and loss of acetylation of histone H3 and H4 were detected. Our results demonstrate that the abnormal transcription and epigenetic modification of sequences flanking the transgene were not correlated with the expression of the transgene. However, the disturbance caused by the insertion of the transgene, was dependent upon the integration site. This suggests that some sequences in the host genome could permit integration and expression of transgene without causing defects in the host.
转基因技术多年来一直被用于研究基因功能、生产重要蛋白质以及生成人类疾病研究模型。然而，产生保留正常功能的转基因动物品系的效率极低。这种低效率主要可归因于整合的转基因。进一步了解转基因整合对宿主基因组转录和表观遗传修饰的影响将提高转基因效率。因此，我们利用通过体细胞核移植产生的三只表达绿色荧光蛋白（GFP）的转基因猪，来鉴定由GFP基因整合导致的转录、DNA甲基化和组蛋白乙酰化的变化。通过热不对称交错PCR（TAIL-PCR）验证了三只转基因猪中转基因多拷贝整合到单个位点，并且每只猪的整合位点不同。我们观察到，与野生型猪相比，整合的转基因经常导致转基因猪各种组织中侧翼序列的转录显著降低。与低转录相对应，检测到DNA高甲基化以及组蛋白H3和H4乙酰化的丧失。我们的结果表明，转基因侧翼序列的异常转录和表观遗传修饰与转基因的表达无关。然而，转基因插入引起的干扰取决于整合位点。这表明宿主基因组中的一些序列可以允许转基因的整合和表达而不会在宿主中造成缺陷。
