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The full length cDNA sequence of the Tpt1/ TCTP (Tumor protein, Translationally-controlled1) gene was identified from Common Carp ( Cyprinus carpio, Cyprinidae), and was designated as CcTpt1 gene. The CDS is 510 bp and encodes a 170-amino acid peptide with a typical Tpt1 signature 2 domain, and is a typical Tpt1 protein. The deduced amino acid sequence of Tpt1 shared significant identity with the Tpt1 from other animals. A phylogenetic tree analysis revealed that the Common Carp Tpt1 protein has the closest genetic relationship and evolutional distance with Tpt1 from Medaka (Oryzias Latipes). Analysis by RT-PCR showed that the Tpt1 mRNA was detected in heart, liver, gill, kidney, muscle and skin. In embryogenesis, the Tpt1 mRNA was expressed gradually stronger from two-cell stage until prim-5 stage by whole-mount in situ. In larval stage, the Tpt1 was specifically expressed at eyes and brain, later at the ear stone, intestines, gills and internal organs. In addition, the Tpt1 was also found to be expressed in skin matrix being developed into scales and gradually disappeared when the scales were fully formed. These data suggested that the CcTpt1 may play important roles in early embryogenesis and scale initiation in fish. [ABSTRACT FROM AUTHOR]

Whole-mount immunofluorescence technique provides a way to reveal integrated expression patterns of biological molecules in individuals. Well-documented morphological preservation ability in biology makes aldehydes the fixative of choice. Cross-linking among biocomponents and aldehydes is the key for maintaining morphology but masks the biological molecules for immunodetection. This study performs an easily accessible method by applying heat-induced retrieval, which can rescue the antigenicity of the proteins and also enhance the labeling sensitivity of the fluorescence dye in overfixed zebrafish embryos. The results show that the immunoreactivities of antibodies to myosin in the muscles, green fluorescent protein in the blood vessels and the nuclei in the cells can be recovered significantly, and the morphology of the zebrafish embryos, even the fragile mutants, is at the same time well maintained. Therefore, we provide a choice for antigen retrieval, which is effective for whole-mount immunofluorescence microscopy. [ABSTRACT FROM AUTHOR]

The root is an ideal model system for studying subcellular localization and dynamic trafficking of important membrane-associated proteins in plants. Immunofluorescence analysis is necessary to reveal subcellular localization and intracellular trafficking of endogenous proteins as epitope tags or fluorescent proteins may cause mislocation of fusion proteins. Here, we describe a rapid and reliable immunodetection protocol for whole-mount in situ localization of membrane-associated proteins involved in clathrin-mediated endocytosis (CME) in Arabidopsis root cells. The whole procedure includes five basic steps: tissue fixation, tissue permeation, blocking, primary antibody incubation, and secondary antibody incubation.