Oxtoby E, Jowett T
Department of Biochemistry and Genetics, Medical School, University of Newcastle upon Tyne, UK.
Nucleic Acids Res. 1993 Mar 11;21(5):1087-95. doi: 10.1093/nar/21.5.1087.
To begin to examine the function of genes that control early development in the hindbrain, we have screened an embryonic zebrafish cDNA library with a murine krox-20 gene probe that contained the conserved zinc finger regions. We have isolated two overlapping cDNAs, zf187 and zf201 which are homologues of the murine krox-20 gene. The N-terminal of the longest cDNA (zf201) contains two acidic regions identical to those of the murine krox-20. This indicates that the functional organisation of these proteins is probably conserved. Northern Blot analysis identified a single transcript of 2.0 kb. Wholemount in situ hybridisation established that expression of the zebrafish gene (krx-20) first appears at 100% epiboly as a single anterior domain of the prospective neuroepithelium, followed very soon after by a second more posterior domain. The alternating pattern of expression of this gene in rhombomeres(r) r3 and r5 is apparent by 12 hr post-fertilisation, that is prior to the morphological appearance of the rhombomeres. Around 14 hr neural crest migration begins from the dorsal surface of r5, moving caudally into r6 and then ventrally towards the pharyngeal arches. Crest migration is not apparent at or after 16 hr. No neural crest migration was observed from r3. Expression of krx-20 is down regulated firstly in r3 around 26 hr and later in r5 around 30 hr.
为了开始研究控制后脑早期发育的基因的功能，我们用含有保守锌指区域的鼠源krox - 20基因探针筛选了一个斑马鱼胚胎cDNA文库。我们分离出了两个重叠的cDNA，zf187和zf201，它们是鼠源krox - 20基因的同源物。最长的cDNA（zf201）的N端包含两个与鼠源krox - 20相同的酸性区域。这表明这些蛋白质的功能结构可能是保守的。Northern印迹分析鉴定出一个2.0 kb的单一转录本。整体原位杂交表明，斑马鱼基因（krx - 20）的表达在原肠胚形成100%时首次出现，作为预期神经上皮的单个前部区域，随后很快出现第二个更靠后的区域。该基因在菱脑节（r）r3和r5中交替表达的模式在受精后12小时就很明显，即在菱脑节的形态出现之前。大约在14小时，神经嵴从r5的背表面开始迁移，向尾侧移入r6，然后向腹侧朝向咽弓。在16小时及之后没有观察到明显的神经嵴迁移。在r3没有观察到神经嵴迁移。krx - 20的表达首先在大约26小时在r3中下调，随后在大约30小时在r5中下调。
