Pineal-specific expression of green fluorescent protein under the control of the serotonin-N-acetyltransferase gene regulatory regions in transgenic zebrafish
Pineal-specific expression of green fluorescent protein under the control of the serotonin-N-acetyltransferase gene regulatory regions in transgenic zebrafish
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DOI：
10.1002/dvdy.10152
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发表时间：
2002-11-01
影响因子：
2.5
通讯作者：
Klein, DC
中科院分区：
文献类型：
--
作者：
Gothilf, Y;Toyama, R;Klein, DC
斑马鱼5-羟色胺-N-乙酰转移酶-2(zfAANAT-2)基因在胚胎阶段的松果体(骨骺)中仅有表达。在这里，我们已经开始努力研究潜在的组织特异性表达该基因的机制。制备了绿色荧光蛋白(GFP)由zfAANAT-2基因调控区驱动的DNA载体。斑马鱼胚胎的体内瞬时表达分析表明，松果体特异性表达除了5‘侧翼区外，还需要X侧翼区的调控序列。这一发现导致了在zfAANAT-2基因的5‘和3’调控区控制下产生表达GFP的转基因株系的努力。将瞬时表达GFP的胚胎培养到成熟，并测试转基因的生殖细胞传播。从受精后1~2天开始检测到松果体中的GFP荧光，共获得3个转基因株系。智力炸弹和浮头突变体分别导致松果体异常发育和zfAANAT-2基因水平的升高或降低。纯合子突变转基因胚胎对松果体中GFP表达的影响相似。这些观察表明，这里描述的转基因株系将有助于研究松果体的发育和决定松果体特异基因在斑马鱼中表达的机制。2002年出版，威利-利斯，lnc。
Zebrafish serotonin-N-acetyltransferase-2 zfAANAT-2) mRNA is exclusively expressed in the pineal gland (epiphysis) at the embryonic stage. Here, we have initiated an effort to study the mechanisms underlying tissue-specific expression of this gene. DNA constructs were prepared in which green fluorescent protein (GFP) is driven by regulatory regions of the zfAANAT-2 gene. In vivo transient expression analysis in zebrafish embryos indicated that in addition to the 5'-flanking region, a regulatory sequence in the X-flanking region is required for pineal-specific expression. This finding led to an effort to produce transgenic lines expressing GFP under the control of the 5' and 3' regulatory regions of the zfAANAT-2 gene. Embryos transiently expressing GFP were raised to maturity and tested for germ cell transmission of the transgene. Three transgenic lines were produced in which GFP fluorescence in the pineal was detected starting 1 to 2 days after fertilization. One line was crossed with mindbomb and floating head mutants that cause abnormal development of the pineal and an elevation or reduction of zfAANAT-2 mRNA levels, respectively. Homozygous mutant transgenic embryos exhibited similar effects on GFP expression in the pineal gland. These observations indicate that the transgenic lines described here will be useful in studying the development of the pineal gland and the mechanisms that determine pineal-specific gene expression in the zebrafish. Published 2002 Wiley-Liss, lnc.