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      Gene Cloning and mRNA Expression of Glutamate Dehydrogenase in the Liver, Brain, and Intestine of the Swamp Eel, Monopterus albus (Zuiew), Exposed to Freshwater, Terrestrial Conditions, Environmental Ammonia, or Salinity Stress

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          Abstract

          The swamp eel, Monopterus albus, is an obligatory air-breathing teleost which can undergo long period of emersion, has high environmental and tissue ammonia tolerance, and can survive in brackish water. We obtained a cDNA sequence of glutamate dehydrogenase ( gdh), which consisted of a 133-bp 5′ UTR, a complete coding sequence region spanning 1629 bp and a 3′ UTR of approximately 717 bp, from the liver, intestine, and brain of M. albus. The translated Gdh amino acid sequence had 542 residues, and it formed a monophyletic clade with Bostrychus sinensis Gdh1a, Tetraodon nigroviridis Gdh1a, Chaenocephalus aceratus Gdh1a, Salmo salar Gdh1a1 and Gdh1a2, and O. mykiss Gdh1a. One day of exposure to terrestrial conditions or 75 mmol l −1 NH 4Cl, but not to water at salinity 20, resulted in a significant increase in mRNA expression of gdh1a and Gdh amination activity in the liver of M. albus. However, exposure to brackish water, but not to terrestrial conditions or 75 mmol l −1 NH 4Cl, led to a significant increase in the mRNA expression of gdh1a and Gdh amination activity in the intestine. By contrast, all the three experimental conditions had no significant effects on the mRNA expression of gdh1a in the brain of M. albus, despite a significant decrease in the Gdh amination activity in the brain of fish exposed to 75 mmol l −1 NH 4Cl for 6 days. Our results indicate for the first time that the mRNA expression of gdh1a was differentially up-regulated in the liver and intestine of M. albus in response to ammonia toxicity and salinity stress, respectively. The increases in mRNA expression of gdh1a and Gdh amination activity would probably lead to an increase in glutamate production in support of increased glutamine synthesis for the purpose of ammonia detoxification or cell volume regulation under these two different environmental conditions.

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          PerlPrimer: cross-platform, graphical primer design for standard, bisulphite and real-time PCR.

          PerlPrimer is a cross-platform graphical user interface application for the design of primers for standard, bisulphite and real-time PCR, and sequencing. The program incorporates highly accurate melting-temperature and primer-dimer prediction algorithms with powerful tools such as sequence retrieval from Ensembl and the ability to BLAST search primer pairs. It aims to automate and simplify the process of primer design. Open-source and freely available from http://perlprimer.sourceforge.net.
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            Comparative genomics provides evidence for an ancient genome duplication event in fish.

            There are approximately 25 000 species in the division Teleostei and most are believed to have arisen during a relatively short period of time ca. 200 Myr ago. The discovery of 'extra' Hox gene clusters in zebrafish (Danio rerio), medaka (Oryzias latipes), and pufferfish (Fugu rubripes), has led to the hypothesis that genome duplication provided the genetic raw material necessary for the teleost radiation. We identified 27 groups of orthologous genes which included one gene from man, mouse and chicken, one or two genes from tetraploid Xenopus and two genes from zebrafish. A genome duplication in the ancestor of teleost fishes is the most parsimonious explanation for the observations that for 15 of these genes, the two zebrafish orthologues are sister sequences in phylogenies that otherwise match the expected organismal tree, the zebrafish gene pairs appear to have been formed at approximately the same time, and are unlinked. Phylogenies of nine genes differ a little from the tree predicted by the fish-specific genome duplication hypothesis: one tree shows a sister sequence relationship for the zebrafish genes but differs slightly from the expected organismal tree and in eight trees, one zebrafish gene is the sister sequence to a clade which includes the second zebrafish gene and orthologues from Xenopus, chicken, mouse and man. For these nine gene trees, deviations from the predictions of the fish-specific genome duplication hypothesis are poorly supported. The two zebrafish orthologues for each of the three remaining genes are tightly linked and are, therefore, unlikely to have been formed during a genome duplication event. We estimated that the unlinked duplicated zebrafish genes are between 300 and 450 Myr. Thus, genome duplication could have provided the genetic raw material for teleost radiation. Alternatively, the loss of different duplicates in different populations (i.e. 'divergent resolution') may have promoted speciation in ancient teleost populations.
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              The zebrafish gene map defines ancestral vertebrate chromosomes.

              Genetic screens in zebrafish (Danio rerio) have identified mutations that define the roles of hundreds of essential vertebrate genes. Genetic maps can link mutant phenotype with gene sequence by providing candidate genes for mutations and polymorphic genetic markers useful in positional cloning projects. Here we report a zebrafish genetic map comprising 4073 polymorphic markers, with more than twice the number of coding sequences localized in previously reported zebrafish genetic maps. We use this map in comparative studies to identify numerous regions of synteny conserved among the genomes of zebrafish, Tetraodon, and human. In addition, we use our map to analyze gene duplication in the zebrafish and Tetraodon genomes. Current evidence suggests that a whole-genome duplication occurred in the teleost lineage after it split from the tetrapod lineage, and that only a subset of the duplicates have been retained in modern teleost genomes. It has been proposed that differential retention of duplicate genes may have facilitated the isolation of nascent species formed during the vast radiation of teleosts. We find that different duplicated genes have been retained in zebrafish and Tetraodon, although similar numbers of duplicates remain in both genomes. Finally, we use comparative mapping data to address the proposal that the common ancestor of vertebrates had a genome consisting of 12 chromosomes. In a three-way comparison between the genomes of zebrafish, Tetraodon, and human, our analysis delineates the gene content for 11 of these 12 proposed ancestral chromosomes.
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                Author and article information

                Journal
                Front Physiol
                Front. Physio.
                Frontiers in Physiology
                Frontiers Research Foundation
                1664-042X
                13 December 2011
                2011
                : 2
                : 100
                Affiliations
                [1] 1simpleDepartment of Biological Science, National University of Singapore Singapore
                [2] 2simpleNatural Sciences and Science Education, National Institute of Education, Nanyang Technological University Singapore
                Author notes

                Edited by: Hans O. Poertner, Alfred Wegener Institute for Polar and Marine Research, Germany

                Reviewed by: Pung P. Hwang, Academia Sinica, Taiwan; J. F. Staples, University of Western Ontario, Canada; Wayne R. Fitzgibbon, Medical University of South Carolina, USA

                *Correspondence: Yuen K. Ip, Department of Biological Sciences, National University of Singapore, Kent Ridge, Singapore 117543. e-mail: dbsipyk@ 123456nus.edu.sg

                This article was submitted to Frontiers in Aquatic Physiology, a specialty of Frontiers in Physiology.

                Article
                10.3389/fphys.2011.00100
                3267175
                22319499
                d532e197-d1d8-4957-89ad-9770fc614eb7
                Copyright © 2011 Tok, Chew and Ip.

                This is an open-access article distributed under the terms of the Creative Commons Attribution Non Commercial License, which permits non-commercial use, distribution, and reproduction in other forums, provided the original authors and source are credited.

                History
                : 06 September 2011
                : 24 November 2011
                Page count
                Figures: 6, Tables: 4, Equations: 0, References: 52, Pages: 11, Words: 8044
                Categories
                Physiology
                Original Research

                Anatomy & Physiology
                ammonia,glutamate,glutamate dehydrogenase,osmoregulation,monopterus albus,mrna expression,nitrogen metabolism,swamp eel

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