Showing papers by "Zhiyuan Gong published in 2003"

Recapitulation of fast skeletal muscle development in zebrafish by transgenic expression of GFP under the mylz2 promoter.

Abstract: A 1,934-bp muscle-specific promoter from the zebrafish mylz2 gene was isolated and characterized by transgenic analysis. By using a series of 5' promoter deletions linked to the green fluorescent protein (gfp) reporter gene, transient transgenic analysis indicated that the strength of promoter activity appeared to correlate to the number of muscle cis-elements in the promoter and that a minimal -77-bp region was sufficient for a relatively strong promoter activity in muscle cells. Stable transgenic lines were obtained from several mylz2-gfp constructs. GFP expression in the 1,934-bp promoter transgenic lines mimicked well the expression pattern of endogenous mylz2 mRNA in both somitic muscle and nonsomitic muscles, including fin, eye, jaw, and gill muscles. An identical pattern of GFP expression, although at a much lower level, was observed from a transgenic line with a shorter 871-bp promoter. Our observation indicates that there is no distinct cis-element for activation of mylz2 in different skeletal muscles. Furthermore, RNA encoding a dominant negative form of cAMP-dependent protein kinase A was injected into mylz2-gfp transgenic embryos and GFP expression was significantly reduced due to an expanded slow muscle development at the expense of GFP-expressing fast muscle. The mylz2-gfp transgene was also transferred into two zebrafish mutants, spadetail and chordino, and several novel phenotypes in muscle development in these mutants were discovered.

Zebrafish atonal homologue zath3 is expressed during neurogenesis in embryonic development.

Abstract: Basic helix-loop-helix (bHLH) transcriptional activators function in development of various cell lineages, including the central nervous system. One of the bHLH proteins, Math3/Xath3/NeuroM, was suggested to act as a late proneural gene in the mouse, Xenopus, and chick. Here, we isolated a zebrafish homologue, named zath3, and analyzed its expression pattern in zebrafish embryos. In the neural plate, zath3 is expressed first in the primordia of the tegmentum and trigeminal ganglia and three classes of primary neurons: sensory neurons, interneurons, and motor neurons. During later development, zath3 transcripts were localized along the boundaries of the optic tectum in the midbrain and rhombomeres of the hindbrain. Analyses of zath3 expression in three mid-hindbrain boundary mutants, acerebellar, no isthmus, and spiel-ohne-grensen, indicated that distribution of zath3 mRNAs in the midbrain and hindbrain was dramatically disturbed. In addition, these mutants also affect expression of zath3 in the neuroretina. Developmental Dynamics 227:587–592, 2003. © 2003 Wiley-Liss, Inc.

Generation of living color transgenic zebrafish

Abstract: Use of green fluorescent protein (gfp) as a reporter gene is a powerful approach for the investigation of tissue-specific gene expression and cellular localization of proteins because the fluorescence of its protein product can be conveniently detected in living cells without supplementing a substrate. The approach is particularly useful in zebrafish because of the transparency and external development of their embryos. In the past few years, using several zebrafish tissue-specific promoters, we have developed several stable lines of gfp transgenic zebrafish that display green fluorescence in different tissues; these include five transgenic lines under an epidermis-specific keratin8 (krt8) promoter, two transgenic lines under a fast skeletal muscle-specific promoter from the myosin light polypeptide 2 (mylz2) gene, and five transgenic lines under an elastaseA (elaA) promoter that is specifically expressed in pancreatic exocrine cells. In all cases, transgenic GFP is faithfully expressed according to the specificity of the promoters used. These gfp transgenic lines are useful for recapitulation of a gene expression program, investigation of tissue and organ development, cell sorting for in vitro cell culture, and construction of cell type-specific cDNA library. Recently, by using two tissue-specific promoters linked to two different reporter genes, gfp and rfp (red fluorescent protein), we have generated two-color transgenic zebrafish that express GFP in skin epidermis and RFP in fast skeletal muscle. Currently, we are also developing gfp transgenic fish for biomonitoring using selected inducible gene promoters that can respond to heavy metals and estrogenic compounds. Thus, generation of living color transgenic zebrafish will have important applications in biotechnology as well as in developmental biology.

Sale of transgenic fish that express gene encoding fluorescent protein

Abstract: Four zebrafish gene promoters, which are skin specific, muscle specific, skeletal muscle specific and ubiquitously expressed respectively, were isolated and ligated to the 5′ end of the EGFP gene. When the resulting chimeric gene constructs were introduced into zebrafish, the transgenic zebrafish emit green fluorescence under a blue light or ultraviolet light according to the specificity of the promoters used. Thus, new varieties of ornamental fish of different fluorescence patterns, e.g., skin fluorescence, muscle fluorescence, skeletal muscle-specific and/or ubiquitous fluorescence, are developed.

Intracellular antifreeze polypeptide and nucleic acids

Abstract: A family of related intracellular skin type antifreeze polypeptides and corresponding coding nucleic acids are provided. These are the first skin type intracellular antifreeze polypeptides and coding nucleic acids ever reported. The polypeptides are naturally expressed in the skin of Winter Flounder, and skin specific promoters are also provided. The polypeptides are used to make cells cold-resistant, and to improve the palatability of cold foods and liquids. Cold resistant eukaryotes and prokaryotes, including plants, animals and bacteria are made using the skin-type intracellular antifreeze polypeptides and nucleic acids.