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1.
Plant vacuolar Na+/H+ antiporter plays an important role in salt tolerance. A vacuolar Na+/H+ antiporter gene TrNHX1 was cloned from Trifolium repens L., a forage legume, by RT-PCR and RACE methods using degenerate oligonucleotide primers. The TrNHX1 sequence contains 2,394 nucleotides and an open-reading frame of 1,626 nucleotides that encodes a protein of 541 amino acids
with a deduced molecular mass of 59.5 kDa. The deduced amino acid sequence of TrNHX1 is 78% identical to that of a vacuolar
Na+/H+ antiporter of Arabidopsis thaliana, AtNHX1, and contains the consensus amiloride-binding domain. TrNHX1 could partially complement the NaCl-sensitive phenotypes
of yeast mutants Δnhx1 and Δena1-4Δnhx1, and a similar complementation was also observed in the presence of LiCl and KCl. In addition, it was found that TrNHX1 suppressed
the hygromycin-sensitive phenotype of yeast mutant Δena1-4Δnhx1. The expression of TrNHX1 in T. repens increased in the presence of 150 mM NaCl, and this result accords with that of Na+ contents determination under the same treatment. These results suggest that TrNHX1 functions as a vacuolar Na+/H+ antiporter and plays an important role in salt tolerance and ion homeostasis in T. repens. 相似文献
2.
Wenjuan Yao Xiaozhao Deng Hui Zhong Miao Liu Pu Zheng Zhihao Sun Yun Zhang 《Journal of industrial microbiology & biotechnology》2009,36(7):911-921
Corynebacterium glutamicum strains are used for the fermentative production of l-glutamate. Five C. glutamicum deletion mutants were isolated by two rounds of selection for homologous recombination and identified by Southern blot analysis.
The growth, glucose consumption and glutamate production of the mutants were analyzed and compared with the wild-type ATCC
13032 strain. Double disruption of dtsR1 (encoding a subunit of acetyl-CoA carboxylase complex) and pyc (encoding pyruvate carboxylase) caused efficient overproduction of l-glutamate in C. glutamicum; production was much higher than that of the wild-type strain and ΔdtsR1 strain under glutamate-inducing conditions. In the absence of any inducing conditions, the amount of glutamate produced by
the double-deletion strain ΔdtsR1Δpyc was more than that of the mutant ΔdtsR1. The activity of phosphoenolpyruvate carboxylase (PEPC) was found to be higher in the ΔdtsR1Δpyc strain than in the ΔdtsR1 strain and the wild-type strain. Therefore, PEPC appears to be an important anaplerotic enzyme for glutamate synthesis in
ΔdtsR1 derivatives. Moreover, this conclusion was confirmed by overexpression of ppc and pyc in the two double-deletion strains (ΔdtsR1Δppc and ΔdtsR1Δpyc), respectively. Based on the data generated in this investigation, we suggest a new method that will improve glutamate production
strains and provide a better understanding of the interaction(s) between the anaplerotic pathway and fatty acid synthesis. 相似文献
3.
Jian-Xia Zhang Kun-Lin Wu Li-Ning Tian Song-Jun Zeng Jun Duan 《Acta Physiologiae Plantarum》2011,33(2):409-417
4.
A genetic transformation system has been developed for callus cells of Crataegus
aronia using Agrobacterium
tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with
5 mg l−1 Indole-3-butyric acid (IBA) and 0.5 mg l−1 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different
types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red
colored callus was obtained on MS medium supplemented with 2 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l−1 kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli
were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l−1 hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this
is the first time to report an Agrobacterium-mediated transformation system in Crataegus
aronia. 相似文献
5.
In our early experiments, a variety of Bupleurum scorzonerifolium-like somatic hybrid plants were obtained from protoplast fusion between Arabidopsis thaliana and UV-treated/untreated B. scorzonerifolium. To compare the effects of UV and γ-ray irradiation on the B. scorzonerifolium partner and obtain Arabidopsis-like hybrids, we designed a novel combination of somatic hybridization between A. thaliana and B. scorzonerifolium. Before protoplast isolation and fusion, the suspension cells of B. scorzonerifolium were irradiated by gamma ray (60Co, 50 Gy with 1.3 Gy min−1). Both parental protoplasts lost regeneration capacity, but over 100 somatic hybrids restored the capacity and developed
to Arabidopsis-like inflorescences and flowers with some characteristics of B. scorzonerifolium. Some hybrid flowers showed yellow sepal, petal, or carpel, whose color was similar to the petal of B. scorzonerifolium; the others had silique of Arabidopsis with angularity of B. scorzonerifolium, and their parts possessed five stamens, the same as B. scorzonerifolium. Cytological analysis showed that three hybrids had Arabidopsis-like karyotypes. Random Amplified Polymorphic DNA (RAPD) and Simple Sequence Repeats (SSR) profiles revealed that both parental
fragments were amplified from these hybrids. These results indicated chromatin introgression from B. scorzonerifolium to A. thaliana, which may be related to the complementation of hybrid inflorescence and flower generation. 相似文献
6.
Deutch CE 《Antonie van Leeuwenhoek》2011,99(4):781-793
Staphylococcus saprophyticus strains ATCC 15305, ATCC 35552, and ATCC 49907 were found to require l-proline but not l-arginine for growth in a defined culture medium. All three strains could utilize l-ornithine as a proline source and contained l-ornithine aminotransferase and Δ1-pyrroline-5-carboxylate reductase activities; strains ATCC 35552 and ATCC 49907 could use l-arginine as a proline source and had l-arginase activity. The proline requirement also could be met by l-prolinamide, l-proline methyl ester, and the dipeptides l-alanyl-l-proline and l-leucyl-l-proline. The bacteria exhibited l-proline degradative activity as measured by the formation of Δ1-pyrroline-5-carboxylate. The specific activity of proline degradation was not affected by addition of l-proline or NaCl but was highest in strain ATCC 49907 after growth in Mueller–Hinton broth. A membrane fraction from this
strain had l-proline dehydrogenase activity as detected both by reaction of Δ1-pyrroline-5-carboxylate with 2-aminobenzaldehyde (0.79 nmol min−1 mg−1) and by the proline-dependent reduction of p-iodonitrotetrazolium (20.1 nmol min−1 mg−1). A soluble fraction from this strain had Δ1-pyrroline-5-carboxylate dehydrogenase activity (88.8 nmol min−1 mg−1) as determined by the NAD+-dependent oxidation of dl-Δ1-pyrroline-5-carboxylate. Addition of l-proline to several culture media did not increase the growth rate or final yield of bacteria but did stimulate growth during
osmotic stress. When grown with l-ornithine as the proline source, S. saprophyticus was most susceptible to the proline analogues L-azetidine-2-carboylate, 3,4-dehydro-dl-proline, dl-thiazolidine-2-carboxylate, and l-thiazolidine-4-carboxylate. These results indicate that proline uptake and metabolism may be a potential target of antimicrobial
therapy for this organism. 相似文献
7.
A novel antifreeze protein cDNA was cloned by RT-PCR from the larva of the yellow mealworm Tenebrio molitor. The coding fragment of 339 bp encodes a protein of 112 amino acid residues and was fused to the expression vectors pET32a
and pTWIN1. The resulted expression plasmids were transformed into Escherischia coli strains BL21 (DE3), ER2566, and Origami B (DE3), respectively. Several strategies were used for expression of the highly
disulfide-bonded β-helix-contained protein with the activity of antifreeze in different expression systems. A protocol for
production of refolded and active T. molitor antifreeze protein in bacteria was obtained. 相似文献
8.
Background
Three mutations in Arabidopsis thaliana strain Columbia – cpr1, snc1, and bal – map to the RPP5 locus, which contains a cluster of disease Resistance genes. The similar phenotypes, gene expression patterns, and genetic interactions observed in these mutants are related to constitutive activation of pathogen defense signaling. However, these mutant alleles respond differently to various conditions. Exposure to mutagens, such as ethyl methanesulfonate (EMS) and γ-irradiation, induce high frequency phenotypic instability of the bal allele. In addition, a fraction of the bal and cpr1 alleles segregated from bal × cpr1 F1 hybrids also show signs of phenotypic instability. To gain more insight into the mechanism of phenotypic instability of the bal and cpr1 mutations, we systematically compared the behavior of these unusual alleles with that of the missense gain-of-function snc1 allele in response to DNA damage or passage through F1 hybrids. 相似文献9.
A fusion gene was constructed by combining the cry1Ac gene of Bacillus thuringiensis strain 4.0718 with a neurotoxin gene, hwtx-1, which was synthesized chemically. In this process, an enterokinase recognition site sequence was inserted in frame between
two genes, and the fusion gene, including the promoter and the terminator of the cry1Ac gene, was cloned into the shuttle vector pHT304 to obtain a new expression vector, pXL43. A 138-kDa fusion protein was mass-expressed
in the recombinant strain XL002, which was generated by transforming pXL43 into B. thuringiensis acrystalliferous strain XBU001. Quantitative analysis indicated that the expressed protein accounted for 61.38% of total
cellular proteins. Under atomic force microscopy, there were some bipyramidal crystals with a size of 1.0 × 2.0 μm. Bioassay
showed that the fusion crystals from recombinant strain XL002 had a higher toxicity than the original Cry1Ac crystal protein
against third-instar larvae of Plutella xylostella, with an LC50 (after 48 h) value of 5.12 μg/mL. The study will enhance the toxicity of B. thuringiensis Cry toxins and set the groundwork for constructing fusion genes of the B. thuringiensis
cry gene and other foreign toxin genes and recombinant strains with high toxicity.
LiQiu Xia and XiaoShan Long contributed equally to this work. 相似文献
10.
11.
Ingo Schmidt 《Current microbiology》2009,59(2):130-138
The ammonia oxidizers Nitrosomonas europaea and Nitrosomonas eutropha are able to grow chemoorganotrophically under anoxic conditions with pyruvate, lactate, acetate, serine, succinate, α-ketoglutarate,
or fructose as substrate and nitrite as terminal electron acceptor. The growth yield of both bacteria is about 3.5 mg protein
(mmol pyruvate)−1 and the maximum growth rates of N. europaea and N. eutropha are 0.094 d−1 and 0.175 d−1, respectively. In the presence of pyruvate and CO2 about 80% of the incorporated carbon derives from pyruvate and about 20% from CO2. Pyruvate is used as energy and only carbon source in the absence of CO2 (chemoorganoheterotrophic growth). CO2 stimulates the chemoorganotrophic growth of both ammonia oxidizers and the expression of ribulose bisphosphate carboxylase/oxygenase
is down-regulated at increasing CO2 concentration. Ammonium, although required as nitrogen source, is inhibitory for the chemoorganotrophic metabolism of N. europaea and N. eutropha. In the presence of ammonium pyruvate consumption and the expression of the genes aceE, ppc, gltA, odhA, and ppsA (energy conservation) as well as nirK, norB, and nsc (denitrification) are reduced. 相似文献
12.
Several matrix-attachment regions (MARs) from animals have been shown to block interactions between an enhancer and promoter
when situated between the two. Since a similar function for plant MARs has not been discerned, we tested the Zea mays
ADH1 5′ MAR, Nicotiana tabacum
Rb7 3′ MAR and a transformation booster sequence (TBS) MAR from Petunia hybrida for their ability to impede enhancer–promoter interactions in Arabidopsis thaliana. Stable transgenic lines containing vectors in which one of the three MAR elements or a 4 kb control sequence were interposed
between the cauliflower mosaic virus
35S enhancer and a flower-specific AGAMOUS second intron-derived promoter (AGIP)::β-glucuronidase (GUS) fusion were assayed for GUS expression in vegetative tissues. We demonstrate that the TBS MAR element, but not the ADH1 or Rb7 MARs, is able to block interactions between the 35S enhancer and AGIP without compromising the function of either with elements from which they are not insulated.
Accession numbers: TBS from Petunia hybrida cultivar V26, GenBank accession number EU864306. 相似文献
13.
14.
Liangliang Li Eric Dion Gabriel Richard Olivier Domingue Martine Jean François J. Belzile 《Plant molecular biology》2009,69(6):675-684
The eukaryotic DNA mismatch repair (MMR) system contributes to maintaining the fidelity of genetic information by correcting
replication errors and preventing illegitimate recombination events. This study aimed to examine the function(s) of the Arabidopsis thaliana PMS1 gene (AtPMS1), one of three homologs of the bacterial MutL gene in plants. Two independent mutant alleles (Atpms1-1 and Atpms1-2) were obtained and one of these (Atpms1-1) was studied in detail. The mutant exhibited a reduction in seed set and a bias against the transmission of the mutant allele.
Somatic recombination, both homologous and homeologous, was examined using a set of reporter constructs. Homologous recombination
remained unchanged in the mutant while homeologous recombination was between 1.7- and 4.8-fold higher than in the wild type.
This increase in homeologous recombination frequency was not correlated with the degree of sequence divergence. In RNAi lines,
a range of increases in homeologous recombination were observed with two lines showing a 3.3-fold and a 3.6-fold increase.
These results indicate that the AtPMS1 gene contributes to an antirecombination activity aimed at restricting recombination between diverged sequences.
Liangliang Li, Eric Dion contributed equally to this work. 相似文献
15.
Besma Sghaier-Hammami Inmaculada Redondo-López Ana M. Maldonado-Alconada Sira Echevarría-Zomeño Jesús V. Jorrín-Novo 《Acta Physiologiae Plantarum》2012,34(3):905-922
The present work is directed at studying changes at the proteome level in Arabidopsis thaliana leaves in response to Pseudomonas syringae virulent (Pst) and avirulent (Pst avrRpt2) strains. Arabidopsis leaves were sampled from challenged plants at 4, 8 and 24 h post inoculation. Proteins were TCA–acetone–phenol
extracted and subjected to 2-DE (5–8 pH range) and MS/MS (MALDI–TOF–TOF) analysis. Out of 800 matched spots on each of the
36 gels analysed, 147 spots were either absent in at least one of the conditions studied (time or treatments; qualitative
variable spots) or differentially accumulated between time and treatments (quantitative variable spots). Out of the 24 proteins
successfully identified over TAIR10 database, 23 have not been reported previously in similar proteomics studies of the Arabidopsis thaliana–Pseudomonas syringae interaction. The exhaustive statistical analysis performed, including principal component and heat map, showed that 24 h
post inoculation can clearly discriminate the challenged plants from the control. The protein change occurred early (4 h post
inoculation) following the virulent pathogen infection, whereas the change occurred later (24 h post inoculation) following
the avirulent pathogen inoculation. Concerning the variable proteins, three behavioural groups can be observed: group 1 (common
protein changes in response to virulent and avirulent pathogen infection), group 2 (protein changes in response to virulent
pathogen infection) and group 3 (protein changes in response to avirulent pathogen infection). Differential identified proteins
following the pathogen infection belonged to different groups including those of oxidative stress defence, enzymes of metabolic
pathways and molecular chaperones. 相似文献
16.
Jing Yu Jiaxi Jiang Zian Fang Yuyang Li Hong Lv Jianping Liu 《Biotechnology letters》2010,32(4):507-512
Inulinase gene (Kcinu) derived from Kluyveromyces cicerisporus was expressed extracellularly in Kluyveromyces lactis using an episomal vector directed by Kcinu promoter. The influence of hap1 gene disruption on the expression of inulinase was studied. Inulinase activity in the supernatant of the recombinant Klhap1Δ strain was 391 U ml−1 after cultured 120 h, which was 2.2-fold that of the wild type host. The relative inulinase mRNA level of the Klhap1Δ strain was 11.3-fold that of the wild type strain, and the expression plasmid was more stable in the mutant host. Based on
these results, the disruption of hap1 facilitated the high and stable expression of inulinase controlled by Kcinu promoter in K. lactis. 相似文献
17.
A chalcone reductase (CHR) gene was isolated from Astragalus membranaceus Bge. var. mongholicus (Bge.) Hsiao (A. mongholicus). The full-length cDNA of A. mongholicus CHR, designated as Amchr (GenBank accession No. HM357239), was 1196 bp long. It had a 957 bp open reading frame encoding a 318-amino acid protein
of 35 kDa, a 67 bp 5′ non-coding region and a 172 bp 3′-untranslated region. The putative AmCHR protein showed striking similarity
to CHR from other leguminous species. Two-dimensional structure modeling showed that AmCHR consisted of 45.28% α-helix, 10.38%
extended strand and 44.34% random coil. Prediction showed that three-dimensional AmCHR was a global protein containing an
aldo-ket-red domain, with a putative Asp-Tyr-Lys-His catalytic tetrad in the center. The AmCHR gene was 1251 bp long, consisting
of three exons and two introns. Intron I was 125 bp and intron II was 169 bp long. Southern blot analysis indicated that Amchr belonged to a small multigene family. Under natural conditions, Amchr was expressed differentially in the root, stem and leaf tissues of A. mongholicus, with a preferential expression in the root. The recombinant AmCHR protein was successfully expressed in Escherichia coli strain BL21 with pET42a vector. The result showed that the expressed AmCHR protein had molecular weight of about 35 kDa,
which matched the size of the predicted protein by bioinformatic analysis. This study opened avenues towards understanding
of the function of AmCHR protein and the role of the Amchr gene in the calycosin-7-O-β-d-glucoside branch pathway in A. mongholicus. 相似文献
18.
Wajahatullah Khan Carlos Costa Alfred Souleimanov Balakrishnan Prithiviraj Donald L. Smith 《Plant Growth Regulation》2011,63(3):243-249
Lipo-chitooligosaccharides (LCOs) are bacteria-to-plant signals required for the establishment of rhizobia–legume nitrogen
fixing symbioses. The ability of LCO [Nod Bj V (C18:1, MeFuc)] isolated from B. japonicum (strain 532C), and of oligomers of chitosan (tetramer, pentamer) and chitin (pentamer) to affect the developmental morphology
of roots in Arabidopsis thaliana (L.) Heynh ecotype Columbia (Col-0) was assessed using an interactive scanner-based image analysis system. LCOs have been
shown to play a role in plant organogenesis at nanomolar concentrations. LCO and the chitin pentamer promoted root growth
and development in Arabidopsis at concentrations of 10 nM and 100 μM, respectively. The LCO treated Arabidopsis plants had about 35% longer roots than untreated control plants. Similarly, treatment with 100 μM chitin pentamer (CHIT5)
resulted in 26% longer roots than the untreated plants; however, chitosan oligomer (CH4 or CH5) treated plants did not differ
from the control plants at either concentration (100 or 1 μM). Both LCOs and the chitin pentamer at higher concentrations
increased root surface area, mean root diameter and number of root tips. However, leaf area increase was observed only in
plants treated with LCO at 10 nM. 相似文献
19.
Bacterial biofilms are associated with chronic infections due to their resistance to antimicrobial agents. Staphylococcus
aureus is a versatile human pathogen and can form biofilms on human tissues and diverse medical devices. To identify novel biofilm
inhibitors of S. aureus, the supernatants from a library of 458 Actinomycetes strains were screened. The culture supernatants (1% v/v) of more than 10 Actinomycetes strains inhibited S. aureus biofilm formation by more than 80% without affecting the growth. The culture supernatants of these biofilm-reducing Actinomycetes strains contained a protease (equivalent to 0.1 μg proteinase K ml−1), which both inhibited S. aureus biofilm formation and detached pre-existing S. aureus biofilms. This study suggests that protease treatment could be a feasible tool to reduce and eradicate S. aureus biofilms. 相似文献
20.
Stefanie Kimbacher Ingrid Gerstl Branko Velimirov Sylvia Hagemann 《Molecular genetics and genomics : MGG》2009,282(2):165-172
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