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In our previous study, we bred a stable cytoplasmic male sterility (CMS) line of tuber mustard by using distant hybridization and subsequent backcrosses. In this CMS plants, all floral organs are normal except the anthers, which are transformed into petals or tubular structures. Recently, 2 mitochondrial genes—atpA and orf220—that are distinctively present in the CMS line of tuber mustard were cloned and partially characterized. In our study of genetic diversity analysis of CMS, 7 species of Brassica and Raphanus crops, which included 5 CMS lines and their respective maintainer lines, were used to compare the constitution of protein-coding genes in the mitochondrial genomes. In 4 of the 43 mitochondrial genes, namely, atpA, orf220, orf256, and orf305/orf324, polymorphisms were detected among the tuber mustard CMS line and its maintainer line. The results of a cluster analysis indicate that petaloid CMS phenotype of tuber mustard is a novel CMS type and is nearer to the nap CMS in Brassica napus at the phylogenetic level. The results of individual amplifications of these genes indicate the presence of 4 sequence-characterized amplified region (SCAR) markers, which enable rapid and reliable identification of this CMS. Expressions of the orf220 and orf256 genes were detected only in the CMS line, while expression of the orf305 gene was detected in the maintainer line. The different expression patterns of different mitochondrial-specific marker genes indicate that the quantity of mitochondrial proteins is differentially regulated during organ/tissue development in tuber mustard. The results of this study suggest that the above mentioned 4 mitochondrial genes are associated with the petaloid CMS phenotype in tuber mustard.  相似文献   

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A novel microbial transglutaminase (TGase) from the cultural filtrate of Streptomyces netropsis BCRC 12429 (Sn) was purified. The specific activity of the purified TGase was 18.2 U/mg protein with an estimated molecular mass of 38 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis. The TGase gene of S. netropsis was cloned and an open reading frame of 1,242 bp encoding a protein of 413 amino acids was identified. The Sn TGase was synthesized as a precursor protein with a preproregion of 82 amino acid residues. The deduced amino acid sequence of the mature S. netropsis TGase shares 78.9–89.6% identities with TGases from Streptomyces spp. A high level of soluble Sn TGase with its N-terminal propeptide fused with thioredoxin was expressed in E. coli. A simple and efficient process was applied to convert the purified recombinant protein into an active enzyme and showed activity equivalent to the authentic mature TGase. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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Liu T  Zhang J  Wang M  Wang Z  Li G  Qu L  Wang G 《Plant cell reports》2007,26(12):2091-2099
DWF4 encodes a rate-limiting mono-oxygenase that mediates 22α-hydroxylation reactions in the BR biosynthetic pathway and it is the target gene in the BR feedback loop. Knockout of DWF4 results in a dwarfed phenotype and other severe defects in Arabidopsis. Here we report on the isolation of the ZmDWF4 gene in maize. Sequence analysis revealed that the open reading frame of ZmDWF4 was 1,518 bp, which encodes a protein composed of 505 amino acid residues with a calculated molecular mass of 57.6 kD and a predicated isoelectric point (pI) of 9.54. Phylogenetic analysis indicated that ZmDWF4 was very close to the Arabidopsis DWF4. In young maize seedlings, the expression of ZmDWF4 in shoots was much higher than that in roots. The highest expression of ZmDWF4 was observed in husk leaves and the lowest in silks during flowering stage. The expression of ZmDWF4 in maize was significantly down regulated by exogenous brassinolide. A heterogeneous complementary experiment demonstrated that the defects of three Arabidopsis DWF4 mutants could be rescued by constitutive expression of ZmDWF4, with leaf expandability, inflorescence stem heights and fertile capabilities all restored to normal levels. Increases in seed and branch number as well as the height of florescence stem were observed in the over-expressed transformants. These findings suggest that ZmDWF4 may be an ortholog gene of Arabidopsis DWF4 and responsible for BR biosynthesis in maize. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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Genes encoding dextranolytic enzymes were isolated from Paenibacillus strains Dex40-8 and Dex50-2. Single, similar but non-identical dex1 genes were isolated from each strain, and a more divergent dex2 gene was isolated from strain Dex50-2. The protein deduced from the Dex40-8 dex1 gene sequence had 716 amino acids, with a predicted Mr of 80.8 kDa. The proteins deduced from the Dex50-2 dex1 and dex2 gene sequences had 905 and 596 amino acids, with predicted Mr of 100.1 kDa and 68.3 kDa, respectively. The deduced amino acid sequences of all three dextranolytic proteins had similarity to family 66 glycosyl hydrolases and were predicted to possess cleavable N-terminal signal peptides. Homology searches suggest that the Dex40-8 and Dex50-2 Dex1 proteins have one and two copies, respectively, of a carbohydrate-binding module similar to CBM_4_9 (pfam02018.11). The Dex50-2 Dex2 deduced amino acid sequence had highest sequence similarity to thermotolerant dextranases from thermophilic Paenibacillus strains, while the Dex40-8 and Dex50-2 Dex1 deduced protein sequences formed a distinct sequence clade among the family 66 proteins. Examination of seven Paenibacillus strains, using a polymerase chain reaction-based assay, indicated that multiple family 66 genes are common within this genus. The three recombinant proteins expressed in Escherichia coli possessed dextranolytic activity and were able to convert ethanol-insoluble blue dextran into an ethanol-soluble product, indicating they are endodextranases (EC 3.2.1.11). The reaction catalysed by each enzyme had a distinct temperature and pH dependence.  相似文献   

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Genome sequence analysis of Xanthomonas oryzae pv. oryzae has revealed a cluster of 12 ORFs that are closely related to the gum gene cluster of Xanthomonas campestris pv. campestris. The gum gene cluster of X. oryzae encodes proteins involved in xanthan production; however, there is little experimental evidence supporting this. In this study, biochemical analyses of xanthan produced by a defined set of X. oryzae gum mutant strains allowed us to preliminarily assign functions to most of the gum gene products: biosynthesis of the pentasaccharide repeating unit for GumD, GumM, GumH, GumK, and GumI, xanthan polymerization and transport for GumB, GumC, GumE, and GumJ, and modification of the pentasaccharide repeating unit for GumF, GumG, and GumL. In addition, we found that the exopolysaccharides are essential but not specific for the virulence of X. oryzae. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Sang-Yoon Kim and Jeong-Gu Kim contributed equally to this work.  相似文献   

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Phosphoinositide-specific phospholipase C (PI-PLC) is an important enzyme, which is a key player involved in eukaryotic signal transduction pathways. In plants, it plays a key role in growth and development as well as environmental stress. However, little is known about its roles in signal transduction during sexual reproduction process. In this study, we cloned and characterized a gene of full-length PI-PLC from ovules of Torenia fournieri, designated as TfPLC1. It was 2,171 bp in length, including an open reading frame encoding a polypeptide of 583 amino acids with molecular mass of 66.02 kDa. The amino acid sequence deduced from the cDNA sequence shows 40–76% similarity to other plant PI-PLCs and contains the characteristic X, Y and C2 domains. Northern blot analysis demonstrated it was predominantly expressed in ovules and flowers. Furthermore, TfPLC1 promoter::GUS transgenic analysis indicated it specifically expressed in ovule, stigma and mature pollen grain. Immunohistochemical staining showed that, in mature stigma, TfPLC1 protein was principally localized in the cells of stigmatic receptive surface. Together, our data suggest that TfPLC1 may play an important role in plant sexual reproduction. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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Monosaccharide transporter (MST) genes of Lupinus polyphyllus and L. albus were cloned, expressed and characterised. The isolation and functional characterisation of a cDNA clone and its corresponding genomic clone of a sugar transporter from L. polyphyllus (LpSTP1) is reported. Phylogenetic comparison of the nucleic and amino acid sequences showed the highest similarity to the AtSTP1 gene from Arabidopsis thaliana, which encodes a high affinity sugar transporter. The similar topology as well as the substrate specificity and expression pattern of LpSTP1 encoded protein additionally support the high similarity to the AtSTP1 gene product. The 1,590 bp LpSTP1 cDNA clone was heterologously expressed in yeast resulting in a fully functional specific sugar transporter. This transformation restored the viability of a yeast deletion mutant, which is devoid of all intrinsic MSTs and thus unable to take up and grow on hexose-containing media. The LpSTP1 protein is postulated to be a high-affinity MST since it supported growth best on media containing 0.2% hexose. Tissue-specific expression of LaSTP1 in L. albus was assayed by real-time PCR, which revealed that the lupin STP1 is mainly expressed in flower buds, flowers and young leaves. The results suggest that the main role of LaSTP1 is to catalyse monosaccharide import in sink tissues to meet increased carbohydrate demand during plant development. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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The gene encoding sucrose phosphorylase (742sp) in Leuconostoc mesenteroides NRRL B-742 was cloned and expressed in Escherichia coli. The nucleotide sequence of the transformed 742sp comprised an ORF of 1,458 bp giving a protein with calculated molecular mass of 55.3 kDa. 742SPase contains a C-terminal amino acid sequence that is significantly different from those of other Leu. mesenteroides SPases. The purified 742SPase had a specific activity of 1.8 U/mg with a K m of 3 mM with sucrose as a substrate; optimum activity was at 37°C and pH 6.7. The purified 742SPase transferred the glucosyl moiety of sucrose to cytosine monophosphate (CMP). Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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Three types of respiratory deficient mitochondrial strains have been reported in Chlamydomonas reinhardtii: a deficiency due to (i) two base substitutions causing an amino acid change in the apocytochrome b (COB) gene (i.e., strain named dum-15), (ii) one base deletion in the COXI gene (dum-19), or (iii) a large deletion extending from the left terminus of the genome to somewhere in the COB gene (dum-1, -14, and -16). We found that these respiratory deficient strains of C. reinhardtii can be divided into two groups: strains that are constantly transformable and those could not be transformed in our experiments. All transformable mitochondrial strains were limited to the type that has a large deletion in the left arm of the genome. For these mitochondria, transformation was successful not only with purified intact mitochondrial genomes but also with DNA-constructs containing the compensating regions. In comparison, mitochondria of all the non-transformable strains have both of their genome termini intact, leading us to speculate that mitochondria lacking their left genome terminus have unstable genomes and might have a higher potential for recombination. Analysis of mitochondrial gene organization in the resulting respiratory active transformants was performed by DNA sequencing and restriction enzyme digestion. Such analysis showed that homologous recombination occurred at various regions between the mitochondrial genome and the artificial DNA-constructs. Further analysis by Southern hybridization showed that the wild-type genome rapidly replaces the respiratory deficient monomer and dimer mitochondrial genomes, while the E. coli vector region of the artificial DNA-construct likely does not remain in the mitochondria.  相似文献   

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Control of organ size is the product of coordinated cell division and expansion. In plants where one of these pathways is perturbed, organ size is often unaffected as compensation mechanisms are brought into play. The number of founder cells in organ primordia, dividing cells, and the period of cell proliferation determine cell number in lateral organs. We have identified the Antirrhinum FORMOSA (FO) gene as a specific regulator of floral size. Analysis of cell size and number in the fo mutant, which has increased flower size, indicates that FO is an organ-specific inhibitor of cell division and activator of cell expansion. Increased cell number in fo floral organs correlated with upregulation of genes involved in the cell cycle. In Arabidopsis the AINTEGUMENTA (ANT) gene promotes cell division. In the fo mutant increased cell number also correlates with upregulation of an Antirrhinum ANT-like gene (Am-ANT) in inflorescences that is very closely related to ANT and shares a similar expression pattern, suggesting that they may be functional equivalents. Increased cell proliferation is thought to be compensated for by reduced cell expansion to maintain organ size. In Arabidopsis petal cell expansion is inhibited by the BIGPETAL (BPE) gene, and in the fo mutant reduced cell size corresponded to upregulation of an Antirrhinum BPE-like gene (Am-BPE). Our data suggest that FO inhibits cell proliferation by negatively regulating Am-ANT, and acts upstream of Am-BPE to coordinate floral organ size. This demonstrates that organ size is modulated by the organ-specific control of both general and local gene networks. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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The MADS box genes participate in different steps of vegetative and reproductive plant development, including the most important phases of the reproductive process. Here we describe the isolation and characterisation of two Asparagus officinalis MADS box genes, AOM3 and AOM4. The deduced AOM3 protein shows the highest degree of similarity with ZAG3 and ZAG5 of maize, OsMADS6 of rice and AGL6 of Arabidopsis thaliana. The deduced AOM4 protein shows the highest degree of similarity with AOM1 of asparagus, the SEP proteins of Arabidopsis and the rice proteins OsMADS8, OsMADS45 and OsMADS7. The high level of identity between AOM1 and AOM4 made impossible the preparation of probes specific for one single gene, so the hybridisation signal previously described for AOM1 is probably due to the expression of both genes. The expression profile of AOM3 and AOM1/AOM4 during flower development is identical, and similar to that of the SEP genes. Asparagus genes, however, are expressed not only in flower organs, but also in the different meristem present on the apical region of the shoot during the flowering season: the apical meristem and the three lateral meristems emerging from the leaf axillary region that will give rise to flowers and lateral inflorescences during flowering season, and to phylloclades and branches during the subsequent vegetative phase. The expression of AOM3 and AOM1/AOM4 in these meristems appears to be correlated with the reproductive function of the apex as the hybridisation signal disappears when the apex switches to vegetative function.  相似文献   

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P transposons belong to the eukaryotic DNA transposons, which are transposed by a cut and paste mechanism using a P-element-coded transposase. They have been detected in Drosophila, and reside as single copies and stable homologous sequences in many vertebrate species. We present the P elements Pcin1, Pcin2 and Pcin3 from Ciona intestinalis, a species of the most primitive chordates, and compare them with those from Ciona savignyi. They showed typical DNA transposon structures, namely terminal inverted repeats and target site duplications. The coding region of Pcin1 consisted of 13 small exons that could be translated into a P-transposon-homologous protein. C. intestinalis and C. savignyi displayed nearly the same phenotype. However, their P elements were highly divergent and the assumed P transposase from C. intestinalis was more closely related to the transposase from Drosophila melanogaster than to the transposase of C. savignyi. The present study showed that P elements with typical features of transposable DNA elements may be found already at the base of the chordate lineage. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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Achromobacter xylosoxidans is known to utilize d-glucose via the modified Entner-Doudoroff pathway. Although d-gluconate dehydratase produced from this bacterium was purified and partially characterized previously, a gene that encodes this enzyme has not yet been identified. To obtain protein information on bacterial d-gluconate dehydratase, we partially purified d-gluconate dehydratase in A. xylosoxidans and investigated its biochemical properties. Two degenerate primers were designed based on the N-terminal amino acid sequence of the partially purified d-gluconate dehydratase. Through PCR performed using degenerate primers, a 1,782-bp DNA sequence encoding the A. xylosoxidans d-gluconate dehydratase (gnaD) was obtained. The deduced amino acid sequence of A. xylosoxidans gnaD showed strong similarity with that of proteins belonging to the dihydroxy-acid dehydratase/phosphogluconate dehydratase family (COG0129). This is in contrast to the archaeal d-gluconate dehydratase, which belongs to the enolase superfamily (COG4948). The phylogenetic tree showed that A. xylosoxidans d-gluconate dehydratase is closer to the 6-phosphogluconate dehydratase than the dihydroxy-acid dehydratase. Interestingly, a clade containing A. xylosoxidans enzyme was clustered with proteins annotated as a second and a third dihydroxy-acid dehydratase in the genomes of Clostridium acetobutylicum (Cac_ilvD2) and Streptomyces ceolicolor (Sco_ilvD2, Sco_ilvD3), indicating that the function of these enzymes is the dehydration of d-gluconate.  相似文献   

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The nucleotide sequences of ten SP11 and nine SRK alleles in Raphanus sativus were determined, and deduced amino acid sequences were compared with those of Brassica SP11 and SRK. The amino acid sequence identity of class-I SP11s in R. sativus was about 30% on average, the highest being 52.2%, while that of the S domain of class-I SRK was 77.0% on average and ranged from 70.8% to 83.9%. These values were comparable to those of SP11 and SRK in Brassica oleracea and B. rapa. SP11 of R. sativus S-21 was found to be highly similar to SP11 of B. rapa S-9 (89.5% amino acid identity), and SRK of R. sativus S-21 was similar to SRK of B. rapa S-9 (91.0%). SP11 and SRK of R. sativus S-19 were also similar to SP11 and SRK of B. oleracea S-20, respectively. These similarities of both SP11 and SRK alleles between R. sativus and Brassica suggest that these S haplotype pairs originated from the same ancestral S haplotypes.  相似文献   

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In vertebrate development the Dickkopf protein family carries out multiple functions and is represented by at least four different genes with distinct biological activities. In invertebrates such as Drosophila and Caenorhabditis, Dickkopf genes have so far not been identified. Here we describe the identification and characterization of a Dickkopf gene with a deduced amino acid sequence closely related to that of chicken Dkk-3 in the basal metazoan Hydra. HyDkk-3 appears to be the only Dickkopf gene in Hydra. The gene is expressed in the gastric region in nematocytes at a late differentiation stage. In silico searches of EST and genome databases indicated the absence of Dkk genes from the protostomes Drosophila and Caenorhabditis, whereas within the deuterostomes, a Dkk-3 gene could be identified in the genome of the urochordate Ciona intestinalis. The results indicate that at an early stage of evolution of multicellularity Dickkopf proteins have already played important roles as developmental signals. They also suggest that vertebrate Dkk-1, 2 and 4 may have originated from a common ancestor gene of Dkk-3.H. Fedders and R. Augustin contributed equally to this workEdited by D. Tautz  相似文献   

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