Purpose:

Interferon regulatory factor 4 (IRF4) is identified as a transcriptional factor and plays an important role in the immune response in mammals; however, there are few reports about the function of zebrafish IRF4.

Methods:

We first amplified the coding sequence of irf4a from the testis of zebrafish. Besides, the fragments of irf4a, P2A, EGFP, and Tol2 vector were added for homologous recombination. By sequencing, we can get the Tol2-ef1α-irf4a-EGFP recombinant plasmid and it was microinjected into zebrafish embryos. Fluorescence observation was proceeded at days 3 post fertilization; F0 generations expressing green fluorescence in multiple tissues throughout the body were screened as the founder and raised them to sexual maturity. After mating with WT zebrafish to generate F1 offspring, polymerase chain reaction was used to identify whether irf4a was integrated into the zebrafish genome.

Conclusion:

We obtained the systematic overexpressed irf4a transgenic zebrafish with green fluorescence labeled in spine, eyes, heart, brain, and other tissues. The transgenic zebrafish will be used as a tool for the role of IRF4a in the immune response to the inflammation preconditioning in the future study.

中文翻译：

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Tol2系统构建irf4a转基因斑马鱼及其潜在应用。

目的：

干扰素调节因子 4 (IRF4) 被确定为一种转录因子，在哺乳动物的免疫反应中起重要作用；然而，关于斑马鱼IRF4功能的报道很少。

方法：

我们首先从斑马鱼的睾丸中扩增了irf4a的编码序列。此外，还添加了irf4a、P2A、EGFP和Tol2载体的片段进行同源重组。通过测序，我们可以得到Tol2-ef1α-irf4a-EGFP重组质粒，并将其显微注射到斑马鱼胚胎中。受精后第3天进行荧光观察；在全身多个组织中表达绿色荧光的F0代被筛选为创始人，并将其培养至性成熟。与WT斑马鱼交配产生F1后代后，采用聚合酶链反应鉴定irf4a是否被整合到斑马鱼基因组中。

结论：

我们获得了系统性过表达的irf4a转基因斑马鱼，在脊柱、眼睛、心脏、大脑和其他组织中标记了绿色荧光。在未来的研究中，转基因斑马鱼将用作 IRF4a 在对炎症预处理的免疫反应中的作用的工具。