摘要
研究胚胎发育新基因及其表达规律是揭示胚胎发育基因调控机理的重要途径。我们从18.5天小鼠胚胎脑的cDNA文库中克隆了一个330 bp的cDNA片段,经NCBI检索与0610038D11Rik基因(Genbank登录号为NM_026306)具有完全相同性。该基因定位在小鼠19号染色体E1带,基因组全长1325 bp,包含5个外显子,编码125个氨基酸残基蛋白,含有一个Trm112p蛋白结构域,该基因功能未曾报道。本研究采用RT-PCR,原位杂交和Northern blot对该基因进行表达谱分析,细胞免疫染色对其进行细胞结构定位。全胚胎原位杂交结果显示0610038D11Rik在胚胎E9.5(embryonic day)的端脑,间脑,菱脑和听泡处有较强的信号。随着神经管逐渐关闭,胚胎E10.5在背部神经嵴,神经管区也出现表达信号。E11.5时除了在上述部位表达外,心脏部位也检测到较弱的信号。组织切片原位杂交结果显示在胚胎E12.5～E16基因0610038D11Rik的信号普遍表达,除了在脑部,背脊索神经富集外,在肺、肾、肝脏、肠表达明显;在胸腺、颌下腺、舌、肾上腺、骨骼肌肉也有一定表达,而在心脏信号很弱。RT-PCR实验发现该基因在整个小鼠胚胎发育阶段(E8.5~19.5)均有持续性分布;E15.5的半定量验证了原位杂交的结果,其在多种重要脏器广泛表达,脑、肺、肝、肾表达水平较高,这与新生小鼠的Northern blot实验得到的表达情况相似。细胞定位实验说明其表达的蛋白主要集中在核内和细胞质中。以上研究结果暗示0610038D11Rik基因在小鼠胚胎脑的发育,神经系统和多种器官的形态发生发挥重要的作用。
Research of new genes underlying embryonic development is important for the molecular mechanism during vertebrate embryonic development. A length of 330bp fragment was cloned from a cDNA library of mouse embryonic brain at E18.5 (embryonic day), and homology with gene 0610038D11Rik available in Genbank database. This 0610038D11Rik gene comprises 5 coding exons spanning about 1325 bp of genomic DNA on chromosome19E1, and encodes a 125-amino-acid protein containing a Trm112p-like protein motif. In this study 0610038D11Rik expression patterns were characterized by RT-PCR, northern blot, in situ hybridization, and cellular localization with immuno-cytochemical staining. Here, we report the expression pattern of the gene at embryonic period. A higher expression level of the 0610038D11Rik mRNA was observed in the brain and nervous system by whole mount in situ hybridization analysis during earlier stage of mouse from E9.5 to E11.5. E12.5~16 on sections, revealed a high level of ubiquitous expression, prominent expression was also found in developing brain, spinal cord, and in several developing organs including the thymus, submandibular gland, lung, liver, intestine, metanephros, and gut. In addition, the level of expression persists during the embryonic development (E8.5~E19.5) was detected by RT-PCR. Furthermore, in some tissues, the expression was persistently higher in brain, lung, liver and kidney at E15.5 by RT-PCR and the similar result was observed at P1 (postnatal 1 day) by northern blot analysis. Cellular localization analysis demonstrated that pEGFP-N1-0610038D11Rik was ubiquitous expression into the nucleus and the cytoplasm. Our study suggests that 0610038D11Rik may be implicated in numerous physiological or developmental functions at different embryonic stages. Taken together, these results indicate and serve as a basis for investigating 0610038D11Rik gene product may play a role in the establishment and maintenance of brain development, central nervous system and morphogenesis of diverse tissues and organs.
引文
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