Na^+/K^+-ATPases are membrane-associated enzymes responsible for the active transport of Na^+ and K^+ ions across cell membranes, generating chemical and electrical gradients. These enzymes' α-subunit provides c...Na^+/K^+-ATPases are membrane-associated enzymes responsible for the active transport of Na^+ and K^+ ions across cell membranes, generating chemical and electrical gradients. These enzymes' α-subunit provides catalytic function, binding and hydrolyzing ATP, and itself becoming phosphorylated during the transport cycle. In this study, Na^+/K^+-ATPase α-subunit eDNA was cloned from gill tissue of the swimming crab Portunus trituberculatus by reverse-transcription polymerase chain reaction (RT-PCR) and rapid amplification of eDNA end methods. Analysis of the nucleotide sequence revealed that the eDNA had a full-length of 3 833 base pairs (bp), with an open reading frame of 3 120 bp, 5' untranslated region (UTR) of 317 bp, and 3' UTR of 396 bp. The sequence encoded a 1 039 amino acid protein with a predicted molecular weight of 115.57 kDa and with estimated pI of 5.21. It was predicted here to possess all expected features of Na^+/K^+-ATPase members, including eight transmembrane domains, putative ATP-binding site, and phosphorylation site. Comparison of arnino acid sequences showed that the P. tritubereulatus α-subunit possessed an overall identity of 75%-99% to that of other organisms. Phylogenetic analysis revealed that this α-subunit was in the same category as those of crustaceans. Quantitative real-time RT-PCR analysis indicated that this α-subunit's transcript were most highly expressed in gill and lowest in muscle. RT-PCR analysis also revealed that α-subunit expression in crab gill decreased after 2 and 6 h, but increased after 12, 24, 48, and 72 h. In addition, α-subunit expression in hepatopancreas of crab decreased after 2-72 h. These facts indicated that the crab's Na^+/K^+-ATPase α-subunit was potentially involved in the observed acute response to low salinity stress.展开更多
We conducted this study to assess the diversity of bacteria associated with the surfaces of algae based on 16S rDNA sequence analyses.Twelve strains of bacteria were obtained from the surfaces of the following four sp...We conducted this study to assess the diversity of bacteria associated with the surfaces of algae based on 16S rDNA sequence analyses.Twelve strains of bacteria were obtained from the surfaces of the following four species of algae:Gracilaria textorii,Ulva pertusa,Laminaria japonica,and Polysiphonia urceolata.The isolated strains of bacteria can be divided into two groups:Halomonas and Vibrio,in physiology,biochemical characteristics and 16S rDNA sequence analyses.The phylogenetic tree constructed based on 16S rDNA sequences of the isolates shows four obvious clusters,Halomonas venusta,Vibrio tasmaniensis,Vibrio lentus,and Vibrio splendidus.Isolates from the surface of P.urceolata are more abundant and diverse,of which strains P9 and P28 have a 16S rDNA sequence very similar(97.5%-99.8%) to that of V.splendidus.On the contrary,the isolates from the surfaces of G.textorii,U.pertusa and L.japonica are quite simple and distribute on different branches of the phylogenetic tree.In overall,the results of this study indicate that the genetic relationships among the isolates are quite close and display a certain level of host species specificity,and alga-associated bacteria species are algal species specific.展开更多
A sulfate reducing bacteria was isolated from mining sewage of Daqing Oilfield by Hungate anaerobic technology. Physiological-biochemical analysis showed that the strain could utilize polyacrylamide as sole carbon and...A sulfate reducing bacteria was isolated from mining sewage of Daqing Oilfield by Hungate anaerobic technology. Physiological-biochemical analysis showed that the strain could utilize polyacrylamide as sole carbon and nitrogen source. The sequence analysis of 16S rDNA illustrated that the similarity of F8 and Desulfovibrio desulfuricans (AF192153) was 99%, and the similarity sequence of dissimilatory sulfite reductase gene (DSR) cloned from the strain and Desulfovibrio desulfuricans (AF273034) was 98%. Their phylogenitic analysis was basically anastomosed, and thus temporarily named as Desulfovibrio desulfuricans F8. The DSR cloned from F8 strain was 2740 bp in length consisting of three ORF, DSRA, DSRB and DSRD as a single operon (DSRABD) regulated by the same operator. DSRA contained typical conservative box of sulfate—sulfite reducing enzyme (SiteⅠand SiteⅡ), which could bind siroheme and [Fe4S4]. DSRB retained a [Fe4S4] binding site, with an uncomplimentary structure for siroheme binding. There was no conservative box in DSRD. Sequence analysis of DSR will provide a theoretical basis for quantitative detection, metabolic pathway modification through gene engineering, and sulfate reducing bacteria (SRB) suppression.展开更多
基金Supported by the National High Technology Research and Development Program of China(863 Program)(No.2012AA10A409)the National Natural Science Foundation of China(No.41306177)the Special Scientific Research Funds for Central Non-Profit Institutes,Yellow Sea Fisheries Research Institutes(No.20603022013027)
文摘Na^+/K^+-ATPases are membrane-associated enzymes responsible for the active transport of Na^+ and K^+ ions across cell membranes, generating chemical and electrical gradients. These enzymes' α-subunit provides catalytic function, binding and hydrolyzing ATP, and itself becoming phosphorylated during the transport cycle. In this study, Na^+/K^+-ATPase α-subunit eDNA was cloned from gill tissue of the swimming crab Portunus trituberculatus by reverse-transcription polymerase chain reaction (RT-PCR) and rapid amplification of eDNA end methods. Analysis of the nucleotide sequence revealed that the eDNA had a full-length of 3 833 base pairs (bp), with an open reading frame of 3 120 bp, 5' untranslated region (UTR) of 317 bp, and 3' UTR of 396 bp. The sequence encoded a 1 039 amino acid protein with a predicted molecular weight of 115.57 kDa and with estimated pI of 5.21. It was predicted here to possess all expected features of Na^+/K^+-ATPase members, including eight transmembrane domains, putative ATP-binding site, and phosphorylation site. Comparison of arnino acid sequences showed that the P. tritubereulatus α-subunit possessed an overall identity of 75%-99% to that of other organisms. Phylogenetic analysis revealed that this α-subunit was in the same category as those of crustaceans. Quantitative real-time RT-PCR analysis indicated that this α-subunit's transcript were most highly expressed in gill and lowest in muscle. RT-PCR analysis also revealed that α-subunit expression in crab gill decreased after 2 and 6 h, but increased after 12, 24, 48, and 72 h. In addition, α-subunit expression in hepatopancreas of crab decreased after 2-72 h. These facts indicated that the crab's Na^+/K^+-ATPase α-subunit was potentially involved in the observed acute response to low salinity stress.
基金Supported by the National Natural Science Foundation of China (No 40376048)the National Basic Research Program of China (973 Program) (No 2006CB400604)
文摘We conducted this study to assess the diversity of bacteria associated with the surfaces of algae based on 16S rDNA sequence analyses.Twelve strains of bacteria were obtained from the surfaces of the following four species of algae:Gracilaria textorii,Ulva pertusa,Laminaria japonica,and Polysiphonia urceolata.The isolated strains of bacteria can be divided into two groups:Halomonas and Vibrio,in physiology,biochemical characteristics and 16S rDNA sequence analyses.The phylogenetic tree constructed based on 16S rDNA sequences of the isolates shows four obvious clusters,Halomonas venusta,Vibrio tasmaniensis,Vibrio lentus,and Vibrio splendidus.Isolates from the surface of P.urceolata are more abundant and diverse,of which strains P9 and P28 have a 16S rDNA sequence very similar(97.5%-99.8%) to that of V.splendidus.On the contrary,the isolates from the surfaces of G.textorii,U.pertusa and L.japonica are quite simple and distribute on different branches of the phylogenetic tree.In overall,the results of this study indicate that the genetic relationships among the isolates are quite close and display a certain level of host species specificity,and alga-associated bacteria species are algal species specific.
基金Sponsored by the National Basic Research and Development (973) Program of China(Grant No.2004CB418505)
文摘A sulfate reducing bacteria was isolated from mining sewage of Daqing Oilfield by Hungate anaerobic technology. Physiological-biochemical analysis showed that the strain could utilize polyacrylamide as sole carbon and nitrogen source. The sequence analysis of 16S rDNA illustrated that the similarity of F8 and Desulfovibrio desulfuricans (AF192153) was 99%, and the similarity sequence of dissimilatory sulfite reductase gene (DSR) cloned from the strain and Desulfovibrio desulfuricans (AF273034) was 98%. Their phylogenitic analysis was basically anastomosed, and thus temporarily named as Desulfovibrio desulfuricans F8. The DSR cloned from F8 strain was 2740 bp in length consisting of three ORF, DSRA, DSRB and DSRD as a single operon (DSRABD) regulated by the same operator. DSRA contained typical conservative box of sulfate—sulfite reducing enzyme (SiteⅠand SiteⅡ), which could bind siroheme and [Fe4S4]. DSRB retained a [Fe4S4] binding site, with an uncomplimentary structure for siroheme binding. There was no conservative box in DSRD. Sequence analysis of DSR will provide a theoretical basis for quantitative detection, metabolic pathway modification through gene engineering, and sulfate reducing bacteria (SRB) suppression.