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1.
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
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.
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
4.
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
5.
Heike Helmholz Blair D. Johnston Christiane Ruhnau Andreas Prange 《Hydrobiologia》2010,645(1):223-234
Scyphozoan medusae are very successful foragers which occasionally occur in high abundances in boreal waters and may impact
many different groups in the marine ecosystem by means of a variety of toxins. A rainbow trout gill cell line, RTgill-W1,
was tested for its suitability as quantitative indicator of the cytotoxicity of Cyanea capillata and Aurelia aurita; the major scyphozoan species in the North and Baltic seas. Cultures of rainbow trout gill cells were exposed to whole venoms
extracted from fishing tentacles and oral arms at increasing protein concentrations. The venom caused detachment, clumping
and lysis of cells, as well as a drop in vitality, in a dose-dependent manner. Morphological changes in the cells were evident
within 1 h after venom addition. The damage to gill cells was quantified by measuring the metabolic activity of the cells
by means of the fluorescence of resorufin derived from the nonfluorescent substrate, resazurin. In general, a decrease in
the metabolic activity of the cells was detected at a venom (protein) concentration above 2.0 μg ml−1 (corresponding to 0.2 μg 104 cells−1), and a total loss of activity was observed above 40.0 μg ml−1 (corresponding to 4.0 μg 104 cells−1). C. capillata venoms had increased cytotoxic activity as compared to A. aurita venoms at the same concentration. Cnidocyst extracts from oral arms of A. aurita induced an 85% loss of gill cell viability at concentrations of 0.2 μg 104 cells−1, whereas crude venoms from fishing tentacles reduced cell viability by 18% at the same concentration. Gel electrophoresis
of the venoms indicated that these consist of a large number of proteins in a fairly wide size range, from 6 to 200 kDa, including
some that are the same size as those found in cubomedusae. It also appears that larger (i.e., older) medusae have more complex
venoms and, in some cases, more potent venoms than smaller animals. 相似文献
6.
Recently, the gene coding for a new beta-glucuronidase enzyme has been identified and cloned from Streptococcus equi subsp. zooepidemicus. This is another report of a beta-glucuronidase gene cloned from bacterial species. The ORF Finder analysis of a sequenced
DNA (EMBL, AJ890474) revealed a presence of 1,785 bp large ORF potentially coding for a 594 aa protein. Three protein families
in (Pfam) domains were identified using the Conserved Domain Database (CDD) analysis: Pfam 02836, glycosyl hydrolases family
2, triose phosphate isomerase (TIM) barrel domain; Pfam 02837, glycosyl hydrolases family 2, sugar binding domain; and Pfam
00703, glycosyl hydrolases family 2, immunoglobulin-like beta-sandwich domain. To gain more insight into the enzymatic activity,
the domains were used to generate a bootstrapped unrooted distance tree using ClustalX. The calculated distances for two domains,
TIM barrel domain, and sugar-binding domain were comparable and exhibited similarity pattern based on function and thus being
in accordance with recently published works confirming beta-glucuronidase activity of the enzyme. The calculated distances
and the tree arrangement in the case of centrally positioned immonoglobulin-like beta-sandwich domain were somewhat higher
when compared to other two domains but clustering with other beta-glucuronidases was rather clear. Nine proteins, including
beta-glucuronidases, beta-galactosidase, and mannosidase were selected for multiple alignment and subsequent distance tree
creation. 相似文献
7.
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9.
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
10.
Nadiawati Alias Nor Muhammad Mahadi Abdul Munir Abdul Murad Farah Diba Abu Bakar Nik Azmi Nik Mahmood Rosli Md Illias 《World journal of microbiology & biotechnology》2009,25(4):561-572
A gene encoding endochitinase from Trichoderma virens UKM-1 was cloned and expressed in E. coli BL21 (DE3). Both the endochitinase gene and its cDNA sequences were obtained. The endochitinase gene encodes 430 amino acids
from an open reading frame comprising of 1,690 bp nucleotide sequence with three introns. The endochitinase was expressed
as soluble and active enzyme at 20°C when induced with 1 mM IPTG. Maximum activity was observed at 4 h of post-induction time.
SDS-PAGE showed that the purified endochitinase exhibited a single band with molecular weight of 42 kDa. Biochemical characterization
of the enzyme displayed a near neutral pH characteristic with an optimum pH at 6.0 and optimum temperature at 50°C. The enzyme
is stable between pH 3.0–7.0 and is able to retain its activity from 30 to 60°C. The presence of Mg2+ and Ca2+ ions increased the enzyme activity up to 20%. The purified enzyme has a strong affinity towards colloidal chitin and low
effect on ethyl cellulose and D-cellubiose which are non-chitin related substrates. HPLC analysis from the chitin hydrolysis
showed the release of (GlcNAc)3, (GlcNAc)2 and GlcNAc, in which (GlcNAc)2 was the main product. 相似文献
11.
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
12.
We investigated the spatial distribution and taxonomic identity of mycorrhizal fungi colonizing the root systems of two threatened
Cephalanthera species, C. falcata and C. erecta, in naturally regenerated forests. Peloton formation was observed in both plant species, confirming the existence of orchid
mycorrhizas. For C. falcata, mycorrhization was significantly different among individuals, ranging from 14 to 63%, and no significant difference among
C. erecta individuals was detected (57–68%). Mycorrhization among three growth directions of roots and between orchid species was not
significantly different. The spatial distribution of mycorrhizas in both orchids showed significant differences, being most
frequent at an apical position. Based on DNA sequencing and phylogenetic analyses, we inferred that the families Thelephoraceae
and Sebacinaceae were mycobionts for C. falcata and Thelephoraceae for C. erecta. Our findings indicated that mycorrhizal colonization occurs at a distal position from the base of these orchid root systems
and that mycorrhizal fungi are restricted to few ectomycorrhizal fungal families. 相似文献
13.
14.
Ying Xiao Peng Di Junfeng Chen Ying Liu Wansheng Chen Lei Zhang 《Molecular biology reports》2009,36(7):2019-2029
A novel 4-hydroxyphenylpyruvate dioxygenase gene (designated as Smhppd) was cloned from hairy roots of Salvia
miltiorrhiza Bung. The full-length cDNA of Smhppd was 1,736 bp long with an ORF (open reading frame) that putatively encoded a polypeptide of 481 amino acids, with a predicted
molecular mass of 52.54 kDa. The deduced amino acid sequence of the Smhppd gene shared high homology with other known HPPDs. Analysis of Smhppd genomic DNA revealed that it contained two exons and one intron. The analysis of Smhppd promoter region was also presented. Southern-blot analysis revealed that the Smhppd was a low-copy gene in S.
miltiorrhiza. Real-time quantitative PCR analysis indicated that Smhppd was constitutively expressed in roots, stems and leaves of S.
miltiorrhiza, with the high expression in roots. In addition, Smhppd expreession level under different stress condition was also analyzed during the hairy root culture period, including signaling
components for plant defence responses, such as methyl jasmonate and salicylic acid, as well as an abiotic elicitor, Ag+ and a biotic elicitor, yeast extract. This study will enable us to further understand the role Smhppd plays in the synthesis of active pharmaceutical compounds in S.
miltiorrhiza at molecular level. 相似文献
15.
Ayla Sant’Ana da Silva Javier Freddy Molina Ricardo Sposina Sobral Teixeira Luis G. Valdivieso Gelves Elba P. S. Bon Viridiana S. Ferreira-Leitão 《Biotechnology letters》2017,39(11):1717-1723
Objective
Glucose conversion into disaccharides was performed with β-glucosidases from Prunus dulcis (β-Pd), Aspergillus niger (β-An) and A. awamori (β-Aa), in reactions containing initial glucose of 700 and 900 g l?1.Results
The reactions’ time courses were followed regarding glucose and product concentrations. In all cases, there was a predominant formation of gentiobiose over cellobiose and also of oligosaccharides with a higher molecular mass. For reactions containing 700 g glucose l?1, the final substrate conversions were 33, 38, and 23.5% for β-An, β-Aa, and β-Pd, respectively. The use of β-An yielded 103 g gentiobiose l?1 (15.5% yield), which is the highest reported for a fungal β-glucosidase. The increase in glucose concentration to 900 g l?1 resulted in a significant increase in disaccharide synthesis by β-Pd, reaching 128 g gentiobiose l?1 (15% yield), while for β-An and β-Aa, there was a shift toward the synthesis of higher oligosaccharides.Conclusion
β-Pd and the fungal β-An and β-Aa β-glucosidases present quite dissimilar kinetics and selective properties regarding the synthesis of disaccharides; while β-Pd showed the highest productivity for gentiobiose synthesis, β-An presented the highest specificity.16.
Despite the widespread distribution of Astyanax bockmanni in streams from Upper Paraná River system in central, southeastern, and southern Brazil, just recently, it has been identified
as a distinct Astyanax species. Cytogenetic studies were performed in two populations of this species, revealing conservative features. A. bockmanni shows 2n = 50 chromosomes, a karyotypic formula composed of 10 M + 12SM + 12ST + 16A and multiple Ag-NORs. Eight positive signals
in subtelocentric/acrocentric chromosomes were identified by fluorescent in situ hybridization (FISH) with 18S rDNA probes.
After FISH with 5S rDNA probes, four sites were detected, comprising the interstitial region of a metacentric pair and the
terminal region on long arms of another metracentric pair. Little amounts of constitutive heterochromatin were observed, mainly
distributed at distal region in two chromosomal pairs. Additionally, heterochromatin was also located close to the centromeres
in some chromosomes. No positive signals were detected in the chromosomes of A. bockmanni by FISH with the As-51 satellite DNA probe. The studied species combines a set of characteristics previously identified in
two different Astyanax groups. The chromosomal evolution in the genus Astyanax is discussed. 相似文献
17.
Scheibner M Hülsdau B Zelena K Nimtz M de Boer L Berger RG Zorn H 《Applied microbiology and biotechnology》2008,77(6):1241-1250
Two extracellular enzymes (MsP1 and MsP2) capable of efficient β-carotene degradation were purified from culture supernatants
of the basidiomycete Marasmius scorodonius (garlic mushroom). Under native conditions, the enzymes exhibited molecular masses of ~150 and ~120 kDa, respectively. SDS-PAGE
and mass spectrometric data suggested a composition of two identical subunits for both enzymes. Biochemical characterisation
of the purified proteins showed isoelectric points of 3.7 and 3.5, and the presence of heme groups in the active enzymes.
Partial amino acid sequences were derived from N-terminal Edman degradation and from mass spectrometric ab initio sequencing of internal peptides. cDNAs of 1,604 to 1,923 bp, containing open reading frames (ORF) of 508 to 513 amino acids,
respectively, were cloned from a cDNA library of M. scorodonius. These data suggest glycosylation degrees of ~23% for MsP1 and 8% for MsP2. Databank homology searches revealed sequence
homologies of MsP1 and MsP2 to unusual peroxidases of the fungi Thanatephorus cucumeris (DyP) and Termitomyces albuminosus (TAP). 相似文献
18.
Susanne Schlüter Charles M. A. P. Franz Frank Gesellchen Oliver Bertinetti Friedrich W. Herberg Friedrich R. J. Schmidt 《Current microbiology》2009,59(2):206-211
An Enterococcus faecalis mutant strain with a reduced ability for biofilm formation and primary attachment when compared to the high biofilm-forming
wild-type strain was characterized by molecular biological and proteomic approaches. A point mutation in the srt-1 gene, which encodes a sortase-type enzyme and is part of the recently described bee (biofilm enhancer in Enterococcus) gene cluster, could be identified in the mutant strain. The Srt-1 deficiency resulted in a loss of the Bee-2 protein within
a high molecular weight complex in cell surface protein extracts, as determined by mass spectrometry. These findings strongly
suggest a specific linkage of Bee-2 to Bee-1 and Bee-3 within a complex by Srt-1. Furthermore, the identification of specific
pilin motifs conserved in surface proteins of gram-positive bacteria indicated a possible involvement of the bee genes in the formation of pili structures, and may thus play a role in enhancing biofilm formation in Enterococcus faecalis. 相似文献
19.
Federica Taddei Laura Gazza Salvatore Conti Vera Muccilli Salvatore Foti Norberto Edgar Pogna 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2009,119(7):1205-1212
The starch granule proteins from 113 einkorn wheat (Triticum monococcum ssp monococcum) accessions were analyzed by acidic, polyacrylamide gel electrophoresis (A-PAGE), and two-dimensional A-PAGE x SDS-PAGE.
All accessions were confirmed to contain equal amounts of two polypeptide chains corresponding to puroindoline B (Pin-B),
as well as a prominent component plus a faint band corresponding to puroindoline A (Pin-A). When compared with soft-textured
common wheat, “monococcum” accessions showed an increase of 3.2- and 2.7-fold in Pin-A and Pin-B levels on the starch granules,
respectively. In addition, all accessions contained a novel component of the 2S super-family of seed proteins named Einkorn
Trypsin Inhibitor (ETI), which was found to be encoded as a pre-protein 148 residues long. Wild-type ETI encoded by allele
Eti-A
m
1a and “valine-type” ETI encoded by allele Eti-A
m
1b, which occurred in 107 and six einkorn accessions, respectively, were found to accumulate on starch granules as a mature
protein of 121 amino acids with a hydrophobic central domain. The einkorn accessions exhibited an average SKCS index as low
as −2.05 ± 11.4, which is typical of extra-soft kernels. The total surface area of starch granules in “monococcum” wheat,
as determined by visual assessments in counting chambers, was estimated at 764 mm2/mg of starch, and was about 1.5 times higher than that for common wheat. The results are discussed in relation to the identification
of factors that cause the extra-soft texture of einkorn kernels. 相似文献
20.
Özge Özden William M. Ciesla Wayne J. Fuller David J. Hodgson 《Biodiversity and Conservation》2008,17(12):2821-2832
We analysed the influence of contemporary geography on butterfly diversity for islands in the Mediterranean Basin. We found
that island size and distance from the mainland has a significant effect on the number of species. We also used butterflies
as an indicator group to identify the importance of forest habitats for biodiversity conservation in the island of Cyprus.
To understand the relative importance of local vegetation characteristics of butterflies in the Pentadaktylos mountains transect
counts were used to assess the abundance and butterfly diversity in two different forest types. A total of 1,602 butterflies
and 23 species were recorded during this research. We observed highly significant effects of forest type on abundance and
species richness of butterflies. For example, number of butterflies was significantly higher in old forest than young pine
forest. Also, the abundance of endemic butterflies was highest in old forest habitats. Therefore, the survival of the majority
of endemic butterflies in Cyprus may depend on conservation of old forests and their understorey plants. 相似文献