Berg P E, Henderson A, Ripley S, Yu J K, Anderson W F
Biochem Biophys Res Commun. 1983 Nov 15;116(3):959-65. doi: 10.1016/s0006-291x(83)80235-6.
Plasmids were constructed containing the HSV thymidine kinase gene and two copies of X. borealis 5S rDNA. Mouse L TK- cells were transformed with these DNAs, with selection for the TK+ gene. Transformed cells were then analyzed by Southern blot hybridization and hybridization in situ to determine whether integration of the exogenous DNA occurred at regions of chromosomal homology i.e., at the 5S rDNA regions. Four cell lines were analyzed by Southern blots. Differences in restriction endonuclease specificity strongly suggested that integration was at a different site in each cell line. Two cell lines were further analyzed by hybridization in situ; each showed a single integration site, both different from each other and different from the mouse L cell 5S rDNA sites. Therefore, the presence of two copies of the 5S rDNA gene in the DNA introduced by gene transfer and approximately 300-350 copies of the mouse 5S rDNA gene was not sufficient in these experiments to produce homologous integration into a specific site.
构建了含有单纯疱疹病毒胸苷激酶基因和两份北极霞水母5S核糖体DNA的质粒。用这些DNA转化小鼠L TK-细胞，并对TK+基因进行筛选。然后通过Southern印迹杂交和原位杂交分析转化细胞，以确定外源DNA是否整合到染色体同源区域，即5S rDNA区域。通过Southern印迹分析了四个细胞系。限制性内切酶特异性的差异强烈表明每个细胞系中的整合位点不同。对两个细胞系进一步进行原位杂交分析；每个细胞系都显示出一个单一的整合位点，彼此不同，也与小鼠L细胞5S rDNA位点不同。因此，在这些实验中，基因转移引入的DNA中两份5S rDNA基因以及小鼠约300 - 350份5S rDNA基因的存在不足以产生在特定位点的同源整合。
