BACKGROUND Dmrt2a is a zinc finger like transcription factor with several roles during zebrafish early development: left-right asymmetry, synchronisation of the somite clock genes and fast muscle differentiation. Despite the described functions, Dmrt2a mechanism of action is unknown. Therefore, with this work, we propose to identify Dmrt2a downstream genes during zebrafish early development. RESULTS We generated and validated a heat-shock inducible transgenic line, to timely control dmrt2a overexpression, and dmrt2a mutant lines. We characterised dmrt2a overexpression phenotype and verified that it was very similar to the one described after knockdown of this gene, with left-right asymmetry defects and desynchronisation of somite clock genes. Additionally, we identified a new phenotype of somite border malformation. We generated several dmrt2a mutant lines, but we only detected a weak to negligible phenotype. As dmrt2a has a paralog gene, dmrt2b, with similar functions and expression pattern, we evaluated the possibility of redundancy. We found that dmrt2b does not seem to compensate the lack of dmrt2a. Furthermore, we took advantage of one of our mutant lines to confirm dmrt2a morpholino specificity, which was previously shown to be a robust knockdown tool in two independent studies. Using the described genetic tools to perform and validate a microarray, we were able to identify six genes downstream of Dmrt2a: foxj1b, pxdc1b, cxcl12b, etv2, foxc1b and cyp1a. CONCLUSIONS In this work, we generated and validated several genetic tools for dmrt2a and identified six genes downstream of this transcription factor. The identified genes will be crucial to the future understanding of Dmrt2a mechanism of action in zebrafish.

中文翻译：

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使用及时可控的方法鉴定斑马鱼早期发育过程中的Dmrt2a下游基因。

背景技术Dmrt2a是锌指样转录因子，在斑马鱼的早期发育过程中具有多个作用：左右不对称，somite时钟基因的同步和快速的肌肉分化。尽管有描述的功能，但Dmrt2a的作用机理尚不清楚。因此，通过这项工作，我们建议在斑马鱼的早期发育过程中鉴定Dmrt2a下游基因。结果我们生成并验证了热休克诱导型转基因品系，以及时控制dmrt2a过表达和dmrt2a突变株。我们表征了dmrt2a过表达表型，并验证了它与敲除该基因后所描述的非常相似，具有左右不对称缺陷和somite时钟基因的失步。此外，我们确定了somite边界畸形的新表型。我们产生了几个dmrt2a突变株，但我们仅检测到弱到可忽略的表型。由于dmrt2a具有旁系同源基因dmrt2b，具有相似的功能和表达模式，因此我们评估了冗余的可能性。我们发现dmrt2b似乎无法弥补dmrt2a的不足。此外，我们利用我们的一种突变株来确认dmrt2a吗啉代特异性，该特性先前在两项独立研究中被证明是一种可靠的敲除工具。使用描述的遗传工具执行和验证微阵列，我们能够鉴定Dmrt2a下游的六个基因：foxj1b，pxdc1b，cxcl12b，etv2，foxc1b和cyp1a。结论在这项工作中，我们产生并验证了dmrt2a的几种遗传工具，并鉴定了该转录因子下游的六个基因。