Swoboda P, Gal S, Hohn B, Puchta H
Friedrich Miescher-Institut, Basel, Switzerland.
EMBO J. 1994 Jan 15;13(2):484-9. doi: 10.1002/j.1460-2075.1994.tb06283.x.
A system to assay intrachromosomal homologous recombination during the complete life-cycle of a whole higher eukaryote was set up. Arabidopsis thaliana plants were transformed with a recombination substrate carrying a non-selectable and quantitatively detectable marker gene. The recombination substrates contain two overlapping, non-functional deletion mutants of a chimeric beta-glucuronidase (uidA) gene. Upon recombination, as proven by Southern blot analysis, a functional gene is restored and its product can be detected by histochemical staining. Therefore, cells in which recombination events occurred, and their progeny, can be precisely localized in the whole plant. Recombination was observed in all plant organs examined, from the seed stage until the flowering stage of somatic plant development. Meristematic recombination events revealed cell lineage patterns. Overall recombination frequencies typically were in the range 10(-6)-10(-7) events/genome. Recombination frequencies were found to differ in different organs of particular transgenic lines.
建立了一个用于检测整个高等真核生物完整生命周期内染色体同源重组的系统。用携带非选择性且可定量检测标记基因的重组底物转化拟南芥植株。重组底物包含嵌合β-葡萄糖醛酸酶(uidA)基因的两个重叠的、无功能的缺失突变体。经Southern印迹分析证实,重组后恢复了一个功能基因,其产物可通过组织化学染色检测到。因此,发生重组事件的细胞及其后代可以在整个植株中精确定位。在从种子阶段到体细胞植物发育的开花阶段所检查的所有植物器官中均观察到重组。分生组织中的重组事件揭示了细胞谱系模式。总体重组频率通常在10^(-6)-10^(-7)事件/基因组范围内。发现特定转基因系的不同器官中的重组频率有所不同。