Adám A, Bártfai R, Lele Z, Krone P H, Orbán L
Laboratory of Aquatic Molecular Biology, Agricultural Biotechnology Center, Gödöllo, Hungary.
Exp Cell Res. 2000 Apr 10;256(1):282-90. doi: 10.1006/excr.2000.4805.
Heat-inducibility of two reporter constructs expressing lacZ gene under the control of mouse and Xenopus hsp70 promoters was tested in zebrafish (Danio rerio) embryos using a transient expression system. Cells expressing beta-galactosidase were stained blue by histochemical staining and their average number per embryo was used as an indicator of the expression level of the reporter gene. Both constructs were heat-inducible in the embryonic tissues and showed similar heat dependence (increasing expression levels from 35-36 degrees C up to 39 degrees C with an apparent decrease at 40 degrees C), resembling that of the zebrafish hsp70 genes. However, their induction kinetics were different, which might be due to differences in their 5' UTRs. Spatial expression patterns of the two hsp/lacZ constructs and an endogenous hsp70 gene were mostly similar on the RNA level. These results indicate that our approach is applicable for in vivo analysis of the heat-shock response and that exogenous heat-shock promoters may be useful for inducible expression of transgenes in fish.
利用瞬时表达系统，在斑马鱼（Danio rerio）胚胎中测试了在小鼠和非洲爪蟾hsp70启动子控制下表达lacZ基因的两种报告基因构建体的热诱导性。表达β-半乳糖苷酶的细胞通过组织化学染色染成蓝色，每个胚胎中这些细胞的平均数量用作报告基因表达水平的指标。两种构建体在胚胎组织中均具有热诱导性，并表现出相似的热依赖性（表达水平从35 - 36摄氏度升高至39摄氏度，在40摄氏度时明显下降），类似于斑马鱼hsp70基因。然而，它们的诱导动力学不同，这可能是由于其5' UTR存在差异。在RNA水平上，两种hsp/lacZ构建体和内源性hsp70基因的空间表达模式大多相似。这些结果表明，我们的方法适用于热休克反应的体内分析，并且外源性热休克启动子可能有助于在鱼类中诱导转基因表达。
