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1.
In Escherichia coli cellular levels of pppGpp and ppGpp, collectively called (p)ppGpp, are maintained by the products of two genes, relA and spoT. Like E. coli, Vibrio cholerae also possesses relA and spoT genes. Here we show that similar to E. coli, V. cholerae ΔrelA cells can accumulate (p)ppGpp upon carbon starvation but not under amino acid starved condition. Although like in E. coli, the spoT gene function was found to be essential in V. cholerae
relA
+ background, but unlike E. coli, several V. cholerae ΔrelA ΔspoT mutants constructed in this study accumulated (p)ppGpp under glucose starvation. The results suggest a cryptic source of
(p)ppGpp synthesis in V. cholerae, which is induced upon glucose starvation. Again, unlike E. coli ΔrelA ΔspoT mutant (ppGpp0 strain), the V. cholerae ΔrelA ΔspoT mutants showed certain unusual phenotypes, which are (a) resistance towards 3-amino-1,2,4-triazole (AT); (b) growth in nutrient
poor M9 minimal medium; (c) ability to stringently regulate cellular rRNA accumulation under glucose starvation and (d) initial
growth defect in nutrient rich medium. Since these phenotypes of ΔrelA ΔspoT mutants could be reverted back to ΔrelA phenotypes by providing SpoT in trans, it appears that the spoT gene function is crucial in V. cholerae.
Part of this work was presented at the International Symposium on Chemical Biology, Kolkata, India, 7–9 March 2007. 相似文献
2.
The lipase Lip2 of the edible basidiomycete, Pleurotus sapidus, is an extracellular enzyme capable of hydrolysing xanthophyll esters with high efficiency. The gene encoding Lip2 was expressed
in Escherichia coli TOP10 using the gene III signal sequence to accumulate proteins in the periplasmatic space. The heterologous expression under
control of the araBAD promoter led to the high level production of recombinant protein, mainly as inclusion bodies, but partially
in a soluble and active form. A fusion with a C-terminal His tag was used for purification and immunochemical detection of
the target protein. This is the first example of a heterologous expression and periplasmatic accumulation of a catalytically
active lipase from a basidiomycete fungus. 相似文献
3.
Bimal Kumar Ghimire Eun Soo Seong Jung Dae Lim Kweon Heo Myong Jo Kim Ill-Min Chung John A. Juvik Chang Yeon Yu 《Plant Cell, Tissue and Organ Culture》2008,95(3):265-274
Efficient transformation of leaf disc-derived callus of Codonopsis lanceolata was obtained using Agrobacterium tumefaciens strain LBA4404 harboring a binary vector, pYBI121, that carries the neomycin phosphotransferase (npt II) gene as a selectable marker. The green shoots recovered from agroinfected explants on selection medium (containing 0.1 mg/l
α-naphthaleneacetic acid (NAA), 1 mg/l 6-benzylaminopurine (BAP), 100 mg/l kanamycin, and 250 mg/l cefotaxime) were rooted
on Murashige and Skoog (MS) medium supplemented with 2 mg/l IBA and 10 mg/l kanamycin. To optimize the transformation conditions,
several factors were assessed, including the co-cultivation period, the duration of pre- and post-culture in darkness and
light, the kanamycin concentration, and the Agrobacterium densities. We produced transgenic Codonopsis lanceolata overexpressing γ-tocopherol methyltransferase (γ-TMT) by this protocol. Moreover, the α-tocopherol content of the plants was enhanced by the overexpression of this gene.
Bimal Kumar Ghimire and Eun Soo Seong contributed equally to this work. 相似文献
4.
Maksymilian Zienkiewicz Tomasz Krupnik Anna Drożak Wioleta Wasilewska Anna Golke Elżbieta Romanowska 《Plant molecular biology》2018,96(1-2):135-149
Key message
We have successfully produced single-cell colonies of C. merolae mutants, lacking the PsbQ’ subunit in its PSII complex by application of DTA-aided mutant selection. We have investigated the physiological changes in PSII function and structure and proposed a tentative explanation of the function of PsbQ’ subunit in the PSII complex.Abstract
We have improved the selectivity of the Cyanidioschyzon merolae nuclear transformation method by the introduction of diphtheria toxin genes into the transformation vector as an auxiliary selectable marker. The revised method allowed us to obtained single-cell colonies of C. merolae, lacking the gene of the PsbQ’ extrinsic protein. The efficiency of gene replacement was extraordinarily high, allowing for a complete deletion of the gene of interest, without undesirable illegitimate integration events. We have confirmed the absence of PsbQ’ protein at genetic and protein level. We have characterized the physiology of mutant cells and isolated PSII protein complex and concluded that PsbQ’ is involved in nuclear regulation of PSII activity, by influencing several parameters of PSII function. Among these: oxygen evolving activity, partial dissociation of PsbV, regulation of dimerization, downsizing of phycobilisomes rods and regulation of zeaxanthin abundance. The adaptation of cellular physiology appeared to favorite upregulation of PSII and concurrent downregulation of PSI, resulting in an imbalance of energy distribution, decrease of photosynthesis and inhibition of cell proliferation.5.
Aulus EAD Barbosa Érika VS Albuquerque Maria CM Silva Djair SL Souza Osmundo B Oliveira-Neto Arnubio Valencia Thales L Rocha Maria F Grossi-de-Sa 《BMC biotechnology》2010,10(1):44
Background
Coffee is an important crop and is crucial to the economy of many developing countries, generating around US70 billion per year. There are 115 species in the < i > Coffea < /i > genus, but only two, < i > C. arabica < /i > and < i > C. canephora < /i > , are commercially cultivated. Coffee plants are attacked by many pathogens and insect-pests, which affect not only the production of coffee but also its grain quality, reducing the commercial value of the product. The main insect-pest, the coffee berry borer ( < i > Hypotheneumus hampei < /i > ), is responsible for worldwide annual losses of around US70 billion per year. There are 115 species in the Coffea genus, but only two, C. arabica and C. canephora, are commercially cultivated. Coffee plants are attacked by many pathogens and insect-pests, which affect not only the production of coffee but also its grain quality, reducing the commercial value of the product. The main insect-pest, the coffee berry borer (Hypotheneumus hampei), is responsible for worldwide annual losses of around US500 million. The coffee berry borer exclusively damages the coffee berries, and it is mainly controlled by organochlorine insecticides that are both toxic and carcinogenic. Unfortunately, natural resistance in the genus Coffea to H. hampei has not been documented. To overcome these problems, biotechnological strategies can be used to introduce an α-amylase inhibitor gene (α-AI1), which confers resistance against the coffee berry borer insect-pest, into C. arabica plants. 相似文献6.
Abdul Ghaffar Sher Afzal Khan Zahid Mukhtar Muhammad Ibrahim Rajoka Farooq Latif 《Molecular biology reports》2011,38(5):3227-3233
We studied heterologous expression of xylanase 11A gene of Chaetomium thermophilum in Pichia pastoris and characterized the thermostable nature of the purified gene product. For this purpose, the xylanase 11A gene of C. thermophilum was cloned in P. pastoris GS115 under the control of AOX1 promoter. The maximum extracellular activity of recombinant xylanase (xyn698: gene with intron) was 15.6 U ml−1 while that of recombinant without intron (xyn669) was 1.26 U ml−1 after 96 h growth. The gene product was purified apparently to homogeneity level. The optimum temperature of pure recombinant
xylanase activity was 70°C and the enzyme retained its 40.57% activity after incubation at 80°C for 10 min. It exhibited quite
lower demand of activation energy, enthalpy, Gibbs free energy, entropy, and xylan binding energy during substrate hydrolysis
than that required by that of the donor, thus indicating its thermostable nature. pH-dependent catalysis showed that it was
quite stable in a pH range of 5.5–8.5. This revealed that gene was successfully processed in P. pastoris and remained heat stable and may qualify for its potential use in paper and pulp and animal feed applications. 相似文献
7.
TNFalpha and TNFbeta, or linfotoxin (LTalpha), are two molecules playing an important role in inflammation. Their genes map on Chromosome 6, between the HLA class II and class I loci. Polymorphisms in, or near, TNF genes have been associated with susceptibility to several autoimmune diseases. Studies of TNF genes in celiac disease (CD) have presented contradictory results. We have assessed the role of TNFalpha and linfotoxin alpha (TNFbeta) in CD and their relative value as CD markers in addition to the presence of DQ2. The TNFA -308 polymorphism and the polymorphism at the first intron of the LTA gene were typed in CD patients and healthy controls and the results were correlated with the presence of DQ2. Significant differences were found in genotype and allele frequencies for the TNFA and LTA genes between CD patients and controls, with an increase in the presence of the TNFA*2 and LTA*1 alleles in CD patients. These differences increase when DQ2-positive CD patients and DQ2-positive controls are compared. In DQ2-positive individuals, allele 2 (A) in position -308 of the promoter of TNFA and allele 1 (G) of the NcoI RFLP in the first intron of LTA are additional risk markers for CD. 相似文献
8.
Akhilesh Kumar Amrita Chakraborty Srijani Ghanta Sharmila Chattopadhyay 《Plant Cell, Tissue and Organ Culture》2009,96(2):117-126
Morphologically identical transgenic mint (Mentha arvensis L.) with bacterial glutathione synthetase gene has been developed. Transformed plants were obtained by co-cultivation of
leaf disks with Agrobacterium tumefaciens strain LBA 4404 harbouring a binary vector pCAMBIA-CpGS that carried E. coli glutathione synthetase (GS), β-glucuronidase as reporter gene and nptII as selective marker gene for kanamycin resistance. Using a constitutive double CaMV 35S promoter and an rbcS transit peptide, we successfully addressed CpGS to the chloroplasts through pJIT 117 vector. Preculture and the presence of AS in the co-cultivation medium played a significant
role in enhancing transformation frequency. The highest transformation frequency was achieved with MS selection medium supplemented
with 25% coconut water, 1.12 mg l−1 BAP, 0.2 mg l−1 NAA, 50 mg l−1 kanamycin and 125 mg l−1 cefotaxime. Robust rooting of regenerated shoots was obtained in half-strength liquid MS medium containing 0.2 mg l−1 NAA and 50 mg l−1 kanamycin. The presence and expression of transgenes in transgenics (T0) was evidenced by GUS histoenzymatic assay, PCR and RT-PCR analysis of nptII and the gene of interest, i.e., GS of putative transgenic leaves. Chromosomal integration of GS gene was confirmed by Southern blot analysis. Transgenic plants were successfully acclimatized in the greenhouse. An overall
transformation frequency of 15% was achieved in approximately 3 months of time period. These results are discussed in relation
to heavy metal trafficking pathways in higher plants and to the interest of using plastid expression of PCS for biotechnological applications.
Akhilesh Kumar and Amrita Chakraborty contributed equally. 相似文献
9.
Matías Maggi Natalia Damiani Sergio Ruffinengo David De Jong Judith Principal Martín Eguaras 《Experimental & applied acarology》2010,50(3):269-279
We undertook a field study to determine whether comb cell size affects the reproductive behavior of Varroa destructor under natural conditions. We examined the effect of brood cell width on the reproductive behavior of V. destructor in honey bee colonies, under natural conditions. Drone and worker brood combs were sampled from 11 colonies of Apis mellifera. A Pearson correlation test and a Tukey test were used to determine whether mite reproduction rate varied with brood cell
width. Generalized additive model analysis showed that infestation rate increased positively and linearly with the width of
worker and drone cells. The reproduction rate for viable mother mites was 0.96 viable female descendants per original invading
female. No significant correlation was observed between brood cell width and number of offspring of V. destructor. Infertile mother mites were more frequent in narrower brood cells. 相似文献
10.
11.
Root segments from seedlings of Panax ginseng produced adventitious roots directly when cultured on 1/2 MS solid medium lacking NH4NO3 and containing 3.0 mg l−1 IBA. Using this adventitious root formation, we developed rapid and efficient transgenic root formation directly from adventitious
root segments in P. ginseng. Root segments were co-cultivated with Agrobacterium tumefaciens (GV3101) caring β-glucuronidase (GUS) gene. Putative transgenic adventitious roots were formed directly from root segments on medium with 400 mg l−1 cefotaxime and 50 mg l−1 kanamycin. Kanamycin resistant adventitious roots were selected and proliferated as individual lines by subculturing on medium
with 300 mg l−1 cefotaxime and 50 mg l−1 kanamycin at two weeks subculture interval. Frequency of transient and stable expression of GUS gene was enhanced by acetosyringon (50 mg l−1) treatment. Integration of transgene into the plants was confirmed by the X-gluc reaction, PCR and Southern analysis. Production
of transgenic plants was achieved via somatic embryogenesis from the embryogenic callus derived from independent lines of
adventitious roots. The protocol for rapid induction of transgenic adventitious roots directly from adventitious roots can
be applied for a new Agrobacterium tumefaciens-mediated genetic transformation protocol in P. ginseng. 相似文献
12.
13.
L. Gigliarelli L. Lucentini A. Palomba G. Sgaravizzi H. Lancioni L. Lanfaloni P. Willenz E. Gaino F. Panara 《Hydrobiologia》2008,605(1):265-269
The two freshwater sponges Ephydatia fluviatilis and Ephydatia mülleri belong to the widespread Spongillidae family. Their morphological tracts are very similar and can be distinguished mainly
on the basis of their gemmuloscleres. However, when gemmules are absent it is essential to have an unambiguous species attribution
for a population genetic study based on fresh tissues and historical collections. This article reports a simple Polymerase
Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) analysis, applied to DNA extracted from both gemmules and
fresh tissues in order to discriminate between the two congeneric E. fluviatilis and E. mülleri. Such a biomolecular method is based on the discriminative enzymes’ digestion of each of the three amplified fragments 5.8S-ITS2-28S,
D3 domain of the 28S subunit and COI. Two restriction enzymes were tested for a 620–642 bp fragment of 5.8S-ITS2-28S and for
a 342 bp fragment of the D3 domain of the 28S, one restriction enzyme was tested for a 681 bp fragment codifying for COI.
Obtained digestion patterns were diagnostic for each of the two species, providing a relatively simple, fast and cheap method
for species attribution compared to sequencing.
Handling editor: C. Sturmbauer 相似文献
14.
Dana Bernátová 《Biologia》2008,63(2):175-176
The paper brings information on an isolated occurrence and morphological characters of Carex × involuta and C. juncella populations in the Vel’ká Fatra Mts. Their presence has been known neither from the territory of Slovakia nor from the whole
Western Carpathians till now. 相似文献
15.
Staphylococcus aureus and Staphylococcus epidermidis are two of the most significant opportunistic human pathogens, causing medical implant and nosocomial infections worldwide. These bacteria contain surface proteins that play crucial roles in multiple biological processes. It has become apparent that they have evolved a number of unique mechanisms by which they can immobilise proteins on their surface. Notably, a conserved cell membrane-anchored enzyme, sortase A (SrtA), can catalyse the covalent attachment of precursor bacterial cell wall-attached proteins to peptidoglycan. Considering its indispensable role in anchoring substrates to the cell wall and its effects on virulence, SrtA has attracted great attention. In this study, a 549-bp gene was cloned from a pathogenic S. epidermidis strain, YC-1, which shared high identity with srtA from other Staphylococcus spp. A mutant strain, YC-1ΔsrtA, was then constructed by allelic exchange mutagenesis. The direct survival rate assay suggested that YC-1ΔsrtA had a lower survival capacity in healthy mice blood compare with the wild-type strain, indicating that the deletion of srtA affects the virulence and infectious capacity of S. epidermidis YC-1. YC-1ΔsrtA was then administered via intraperitoneal injection and it provided a relative percent survival value of 72.7 % in mice against S. aureus TC-1 challenge. These findings demonstrate the possbility that YC-1ΔsrtA might be used as a live attenuated vaccine to produce cross-protection against S. aureus. 相似文献
16.
Erwinia carotovora subspecies betavasculorum, also known as E. betavasculorum and Pectobacterium betavasculorum, is a soil bacterium that has the capacity to cause root rot necrosis of sugarbeets. The qualitatively different pathogenicity exhibited by the virulent E. carotovora strain and two avirulent strains, a Citrobacter sp. and an Enterobacter cloacae, was examined using digital analysis of photographic evidence of necrosis as well as for carbohydrate, ethane, and ethylene
release compared with uninoculated potato tuber slices. Visual scoring of necrosis was superior to digital analysis of photographs.
The release of carbohydrates and ethane from potato tuber slices inoculated with the soft rot necrosis-causing Erwinia was significantly greater than that of potato tuber slices that had not been inoculated or that had been inoculated with
the nonpathogenic E. cloacae and Citrobacter sp. strains. Interestingly, ethylene production from potato slices left uninoculated or inoculated with the nonpathogenic
Citrobacter strain was 5- to 10-fold higher than with potato slices inoculated with the pathogenic Erwinia strain. These findings suggest that (1) carbohydrate release might be a useful measure of the degree of pathogenesis, or
relative virulence; and that (2) bacterial suppression of ethylene formation may be a critical step in root rot disease formation. 相似文献
17.
Tamura M Togami J Ishiguro K Nakamura N Katsumoto Y Suzuki K Kusumi T Tanaka Y 《Plant cell reports》2003,21(5):459-466
Verbena (Verbena x hybrida), an important floricultural species, was successfully regenerated from stem segments on Murashige and Skoog's basal medium supplemented with thidiazuron and indole-3-acetic acid. A transformation system was developed using cvs. Temari Scarlet, Temari Sakura, Tapien Rose and TP-P2. Agrobacterium tumefaciens strain Agl0 harboring the sGFP gene was infected into stem segments. Transformation efficiency was improved by evaluating and manipulating the age of the plant material, the concentration of kanamycin in the medium during selection, and the length of the culture period in the dark. After 2-3 months of culture on the selection medium, GFP-positive shoots were obtained in all four of the cultivars tested. These shoots were successfully acclimated and set flowers within 2-3 months in a greenhouse. GFP was expressed in all of the organs including the floral parts. Stable genomic transformation was confirmed by Southern blot analysis. No morphological differences were observed between the transformed plants and their host plants. 相似文献
18.
Recombinant Zantedeschia aethiopica agglutinin (ZAA) was expressed in Escherichia coli as N-terminal His-tagged fusion. After induction with isopropylthio-β-d-galactoside (IPTG), the recombinant ZAA was purified by metal-affinity chromatography. The purified ZAA protein was applied
in anti-fungal assay and the result showed that recombinant ZAA had anti-fungal activity towards leaf mold (Fulvia fulva), one of the most serious phytopathogenic fungi causing significant yield loss of crops. This study suggests that ZAA could
be an effective candidate in genetic engineering of plants for the control of leaf mold. 相似文献
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20.
Fang-Fang Ma Mian Wu Ying-Na Liu Xue-Ying Feng Xun-Zong Wu Jian-Qun Chen Bin Wang 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2018,131(2):253-265