Showing papers by "Zhiyuan Gong published in 2001"

Cloning and developmental expression of a family of pleurocidin-like antimicrobial peptides from winter flounder, Pleuronectes americanus (Walbaum).

Abstract: Low molecular weight antimicrobial peptides are an important component of the innate immune system in animals, yet they have not been examined widely in fish. Of particular interest is their expression during development and in response to environmental conditions and disease. Here, we report the isolation of four genomic sequences encoding putative antimicrobial peptides from the winter flounder, Pleuronectes americanus (Walbaum), as well as reverse transcription-PCR products from two tissues that form the first defensive barrier to microbes — skin and intestine. Alignment of the predicted polypeptide sequences shows a conserved hydrophobic signal peptide of 22 amino acids followed by 25 amino acids that are identical (WF2) or homologous to the amino acid sequence of pleurocidin, followed by a conserved acidic portion. Southern hybridisation analysis indicates that related peptides are encoded in the genomes of other flatfish species. Northern and RT-PCR analyses of RNA from multiple tissues show that two of the pleurocidin genes are expressed predominantly in the skin whereas two other genes are expressed mainly in the intestine. RT-PCR assays of total RNA from larvae of different ages provide the first evidence of developmental expression of antimicrobial peptides in fish and indicate that the pleurocidin gene is first expressed at 13 days post-hatch in winter flounder.

Gene expression pattern Expression pattern of two zebrafish genes, cxcr4a and cxcr4b

Abstract: We cloned and mapped two novel zebrafish genes, cxcr4a and cxcr4b, which are closely related to mammalian CXCR4. Expression analysis by reverse transcription-polymerase chain reaction and in situ hybridization demonstrated that these two genes are expressed in most cell lineages known to express Cxcr4 in mammals. These genes are co-expressed in lateral mesoderm and posterior midbrain. The transcripts of cxcr4a were detected in interneurons and endoderm, whereas cxcr4b was specifically expressed in sensory neurons, motoneurons and cerebellum. In the lateral mesoderm, cxcr4b transcripts appeared earlier than those of cxcr4a. Thus, the function of mammalian CXCR4 could be split between the two zebrafish genes. These genes probably derived from the genome duplication event, which occurred during the evolution of teleosts. Similar pairs of Cxcr4 may exist in other species, where genome duplication has occurred. q 2001 Elsevier Science Ireland Ltd. All rights reserved.

Green fluorescent protein (GFP) transgenic fish and their applications

Abstract: The coupling of the GFP reporter system with the optical clarity of embryogenesis in model fish such as zebrafish and medaka is beginning to change the picture of transgenic fish study. Since the advent of first GFP transgenic fish in 1995, GFP transgenic fish technology have been quickly employed in many areas such as analyses of gene expression patterns and tissue/organ development, dissection of promoters/enhancers, cell lineage and axonal pathfinding, cellular localization of protein products, chimeric embryo and nuclear transplantation, cell sorting, etc. The GFP transgenic fish also have the potentials in analysis of upstream regulatory factors, mutagenesis screening and characterization, and promoter/enhancer trap. Our own studies indicate that GFP transgenic fish may become a new source of novel variety of ornamental fish. Efforts are also being made in our laboratory to turn GFP transgenic fish into biomonitoring organisms for surveillance of environmental pollution.

Multiple tissue transformation in adult zebrafish by gene gun bombardment and muscular injection of naked DNA.

Abstract: The efficiency of two direct gene transfer methods, gene gun (or particle bombardment) and intramuscular injection, in transforming adult zebrafish tissues in vivo was examined by a noninvasive approach using green fluorescent protein (GFP) reporter gene driven by the ubiquitously expressed human cytomegalovirus promoter. Particle bombardment of adult zebrafish caused internalization and expression of the plasmid only in the superficial layer such as epithelial cells, pigment cells, endothelial cells, and neurons, whereas direct injection primarily transformed muscle fibers of several bundles near or around the injection site. Expression was also evident in several nonmuscle tissues, such as skin epithelia, pigment cells, blood vessel cells, and neuron-like cells. GFP expression persisted for more than 50 days with both methods. These observations indicate the potential of these methods for functional analysis of tissue-specific promoters, delivery of DNA vaccine, and muscular expression of other useful genes.