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Hemerythrin is a non-heme respiratory protein involved in oxygen storage and transport in invertebrates. In the present study, the hemerythrin cDNA was cloned from Phascolosoma esculenta (denoted as PeHr) by PCR and rapid amplification of cDNA ends approaches. The full-length PeHr consisted of 770 bp containing of a 5′-terminal untranslated region (UTR) of 83 bp, a 3′-terminal UTR of 327 bp, and a coding domain sequence of 360 bp encoding a polypeptide of 120 amino acids with estimated molecular mass of 13.6 kDa and theoretical isoelectric point of 5.78. The expression profiles of PeHr were evaluated by real-time RT-PCR under blood loss stress. The expression level of PeHr was significantly up-regulated from 45 to 48 h, then slightly recovered to its original level. The coding sequence of the PeHr was cloned and expressed in Escherichia coli BL21 (DE3) for antibodies preparation. Western blotting analysis conformed that the generated antibodies could specifically identify not only recombinant product, but also native protein from the total protein extraction. Our results indicated that PeHr might be involved into haemocytes regeneration, and its function roles should be further investigated by the generated antibodies.  相似文献   

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The methionine adenosyltransferase gene (MAT) had been isolated from an economic seaweed Undaria pinnatifida by PCR using degenerate primers. The cDNA was 1,491 bp in length with an open reading frame of 1,194 nucleotides, encoding a deduced protein of 397 amino acids. The protein had a predicted molecular weight of 43.2 kDa, and the isoelectric point was 5.244. The sequence contains a 92 bp 5′-untranslated region (UTR) and a 205 bp 3′-UTR. The methionine adenosyltransferase (MAT) sequence of U. pinnatifida (UpMAT) shared 68–92 % identities with the previous published MAT sequences of other species. Phylogenetic analysis indicated that the phylogenetic relationship of UpMAT with some other seaweeds was closer than with those of higher plants. Under different stress conditions, the relative mRNA expression levels of the MAT of U. pinnatifida (UpMAT) were measured by real-time quantitative PCR, and the results demonstrated that the UpMAT might help to protect the alga against various abiotic stresses.  相似文献   

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Flavin-containing monooxygenase 3 (FMO3) is an important monooxygenase for catalytic oxygenation of many harmful xenobiotics. Mutations in the FMO3 gene have been identified as causing trimethylaminuria in human and fishy off-flavor in cow milk and chicken eggs. In this study, the full-length cDNA sequence of Pekin duck FMO3 gene was cloned, sequenced, and characterized. The full-length cDNA sequence consisted of 1,846 bp and contained a 1,599 bp open-reading frame encoding 532 amino acids. Duck FMO3 gene shared a similar nine exon–eight intron structure with chicken and human. The duck FMO3 putative protein sequence showed high identity with that of chicken (82 %), and relative low identity with those of mammals (61–66 %). We also found that the duck FMO3 gene was dramatically expressed in liver, lung, and kidney compared to that in other tissues in the ducks, indicating the possible roles the FMO3 gene could play in the three tissues. By bidirectional sequencing, we also found one nonsense mutation, 5 nonsynonymous, and 21 synonymous mutations in the coding region of the FMO3 gene in 11 duck breeds and some of them were predicted to be potentially associated with the activities of FMO3 protein.  相似文献   

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We have cloned and sequenced a 1.7 kb macronuclear chromosome encoding the pheromone 4 gene of Euplotes octocarinatus. The sequence of the secreted pheromone is preceded by a 42 amino acid leader peptide, which ends with a lysine residue. The sequence coding for the leader peptide contains information for a putative signal peptide and is interrupted by a 772 bp intron as shown by comparison with a cDNA clone. A 64 bp intron and a 145 bp intron interrupt the sequence coding for the secreted pheromone. The three introns contain typical 5′ and 3′ splice junctions and a putative branch point site. The small introns have a low GC content. The large intron has a GC content similar to that of the pheromone 4 gene exons. The amino acid sequence of pheromone 4, deduced from both the genomic DNA and the cDNA of pheromone 4, shows that the secreted pheromone consists of 85 amino acids. One of its amino acids is encoded by a UGA codon. Since it has been shown for pheromone 3 of E. octocarinatus that UGA is translated as cysteine, it is assumed that the UGA codon encodes cysteine in pheromone 4 as well. The 164 bp noncoding region upstream of the leader peptide is AT-rich and contains an inverted repeat capable of forming a stem-loop structure with a stem of 11 bp. The 151 bp noncoding region at the 3′ end of the chromosome contains a putative polyadenylation sequence and an inverted repeat. The macro-nuclear molecule is flanked by telomeres and carries the pentanucleotide motif TTGAA, located at a distance of 17 nucleotides from the telomeres. This motif has been suggested to be involved in the formation of macronuclear chromosomes. © 1992 Wiley-Liss, Inc.  相似文献   

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Polymorphisms in the FTO gene are associated with obesity and body mass index in humans and livestock. Little information of whether FTO plays an important role in aquaculture fish species is available. We cloned and characterized the FTO gene in an economically important food fish species: Asian seabass (Lates calcarifer). The full-length cDNA of the gene is 3679 bp, containing an ORF of 1935 bp encoding 644 amino acids, a 216 bp 5′ UTR and a 1538 bp 3′ UTR. The gene consisted of nine exons and eight introns and was 117,679 bp in length. Phylogenetic analysis revealed that the gene in Asian seabass was closely related to those of Japanese flounder and Nile tilapia. Analysis of its expressions using qRT-PCR showed that it was expressed ubiquitously, but was higher in the liver, stomach and intestine. Comparative analysis of the genomic sequences of part of intron 1 of the gene among 10 unrelated individuals identified two SNPs. Analysis of associations between SNPs and traits (i.e. growth, oil content, omega-3 and -6 contents) in an F2 family demonstrated that the two SNPs were significantly associated with growth, oil content, omega-3 content and omega-3/-6 ratio. Altogether, our data suggest that the gene or/and its linked genes play an important role in growth and fatty acid synthesis, and that the SNPs associated with traits may be used as markers for selecting quicker growth and higher omega-3/-6 ratio at the fingerling stage.  相似文献   

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Proteoid roots are a unique adaptation that allow white lupin (Lupinus albus L. var Ultra) to survive under extreme phosphorus (P) deficient conditions. The cascade of events that signals P-deficiency induced gene expression in proteoid roots remains unknown. Through promoter::GUS analysis we showed that expression of acid phosphatase (LaSAP1) in P-deficient proteoid roots depends on DNA located from ?465 bp to ?345 bp 5′ of the ATG start codon and that the P1BS (PHR1 Binding Site) element, located at ?160 bp, also contributes regulatory control. DNA located within the ?414 bp to ?250 bp region of the LaSAP1 promoter was bound by nuclear proteins isolated from P-sufficient normal roots in electrophoretic mobility shift assays (EMSA), suggesting negative regulation. Competition experiments were performed with unlabeled oligonucleotides to further delineate the region of the LaSAP1 promoter bound by P-sufficient normal root nuclear proteins to a motif spanning ?361 bp to ?346 bp. The promoter motif characterized through EMSA spanning ?361 bp to ?345 bp was used as “bait” in a yeast one-hybrid (Y1H) experiment and 31 putative DNA binding proteins were isolated. Taken together, our results increase understanding of P-deficiency signaling by identifying regulatory regions and putative regulatory proteins for LaSAP1 expression.  相似文献   

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