Method for detection and identification of multiple chromosomal integration sites in transgenic animals created with lentivirus
Method for detection and identification of multiple chromosomal integration sites in transgenic animals created with lentivirus
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DOI：
10.2144/000112289
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发表时间：
2006-12-01
期刊：
影响因子：
2.7
通讯作者：
Bauer, Beth A.
中科院分区：
文献类型：
--
作者：
Bryda, Elizabeth C.;Pearson, Michael;Bauer, Beth A.
转基因传递系统，特别是那些涉及逆转录病毒的系统，往往导致转基因的多个副本在整个宿主基因组中整合。由于转基因可以发生位点特异性沉默，因此确定转基因的多个拷贝的数量和染色体位置变得很重要，以便将特定染色体位置的转基因遗传与特定和强大的表型相关联。利用一种结合限制性内切酶消化和几轮PCR扩增以及核苷酸测序的技术，有可能在转基因创始动物中识别多个染色体整合位点。通过设计基因分型分析来检测这些创始人后代中的每个个体整合位点，可以有效地跟踪在特定染色体位置整合的转基因的遗传，因为转基因在随后的世代中随机分离。然后，表型特征可以与整合在特定染色体位置的转基因的遗传相关，从而允许合理选择育种动物，以便建立转基因品系。
Transgene delivery systems, particularly those involving retroviruses, often result in the integration of multiple copies of the transgene throughout the host genome. Since site-specific silencing of trangenes can occur, it becomes important to identify the number and chromosomal location of the multiple copies of the transgenes in order to correlate inheritance of the transgene at a particular chromosomal site with a specific and robust phenotype. Using a technique that combines restriction endonuclease digest and several rounds of PCR amplification followed by nucleotide sequencing, it is possible to identify multiple chromosomal integration sites in transgenic founder animals. By designing genotyping assays to detect each individual integration site in the offspring of these founders, the inheritance of transgenes integrated at specific chromosomal locations can be followed efficiently as the transgenes randomly segregate in subsequent generations. Phenotypic characteristics can then be correlated with inheritance of a transgene integrated at a particular chromosomal location to allow rational selection of breeding animals in order to establish the transgenic line.