bluuebunny/crossref_metadata_2025_split huggingface dataset:
Applying target region amplification polymorphism markers for analyzing genetic diversity of Lentinula edodes in China
Authors: Xiao Yang , Liu Wei , Lu Ying‐Ying , Gong Wen‐Bing , Bian Yin‐Bing 
Published: August 2010

AbstractThe target region amplification polymorphism (TRAP) technique was utilized for assessing the genetic diversity of 55 wild strains and one cultivated strain of Lentinula edodes in China. From these strains, 932 DNA fragments were amplified using 12 primer combinations, 929 fragments (99.68%) of which were polymorphic between two or more strains. The average coefficient of pairwise genetic similarity was 0.696, within a range from 0.503 to 0.947. Cluster analysis and principal coordinate analysis separated the tested strains of L. edodes into two major groups. Group A was further divided into seven subgroups. In most cases, the strains from the same or adjoining regions could be preferentially clustered into small groups. The results from the average genetic similarity and the weighted average value of Shannon's Information Index among the tested strains of L. edodes from the same region revealed a vast genetic diversity in the natural germplasm found in China. Compared with the L. edodes strains from other regions, those found on the Yunnan Plateau, in the Hengduanshan Mountains, in Taiwan, South China, and Northeast China showed greater genetic diversity. The results of the present study indicated that the wild strains of L. edodes in China possessed abundant genetic variation, and the genetic relationships among them were highly associated with the geographic distribution. This is the first report demonstrating that TRAP markers were powerful for analyzing the genetic diversity of L. edodes, and the study lays the foundation for a further application of this remarkable technique to other fungi. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Optimization of a genome‐walking method to suit GC‐rich template DNA from biotechnological relevant Actinobacteria
Authors: Gröning Janosch A. D. , Tischler Dirk , Kaschabek Stefan R. , Schlömann Michael 
Published: August 2010

AbstractSiteFinding‐PCR has been recently reported to be a useful technique in order to identify unknown DNA fragments located adjacent to available sequences. However, this method has so far only been applied to few DNA sources including plants, samples from bioleaching communities, and a Pseudomonas strain. In order to complete the sequence information of two gene clusters in Gram‐positive rhodococci the original protocol was applied yielding amplicons of insufficient size. The binding site of the previously published SiteFinder‐2 oligo proved to be unsuitable for Rhodococcus and other members of the Actinobacteria since the binding motif occurred too frequently. Available genome sequences of different Actinobacteria were analysed and the binding site of the SiteFinder oligo modified. Moreover, PCR conditions were adapted to the high GC content of the template DNA allowing the successful adaptation of this method to two members of the Actinobacteria. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Purification and characterization of the ncgl2923 ‐encoded 3‐hydroxybenzoate 6‐hydroxylase from Corynebacterium glutamicum
Authors: Yang Yi‐Fan , Zhang Jun‐Jie , Wang Song‐He , Zhou Ning‐Yi 
Published: August 2010

AbstractCorynebacterium glutamicum ATCC 13032 metabolizes 3‐hydroxybenzoate via gentisate. We have now characterized the ncgl2923 ‐encoded 3‐hydroxybenzoate 6‐hydroxylase involved in the initial step of 3‐hydroxybenzoate catabolism by this strain, a first 3‐hydroxybenzoate 6‐hydroxylase molecularly and biochemically characterized from a Gram‐positive strain. The ncg12923 gene from Corynebacterium glutamicum ATCC 13032 was shown to encode 3‐hydroxybenzoate 6‐hydroxylase, the enzyme that catalyzes the NADH‐dependent conversion of 3‐hydroxybenzoate to gentisate. Ncgl2923 was expressed with an N‐terminal six‐His tag and purified to apparent homogeneity by Ni2+‐nitrilotriacetic acid affinity chromatography. The purified H6‐Ncgl2923 showed a single band at apparent molecular mass of 49 kDa on a sodium dodecyl sulfate polyacrylamide gel electrophoresis and was found to be most likely a trimer as determined by gel filtration chromatography. It had a specific activity of 6.92 ± 0.39 U mg–1 against 3‐hydroxybenzoate and with a Km value of 53.4 ± 4.7 μM using NADH as a cofactor. The product formed from the 3‐hydroxybenzoate hydroxylation catalyzed by H6‐Ncgl2923 was identified by high‐performance liquid chromatography as gentisate, a ring‐cleavage substrate in the microbial aromatic degradation. The enzyme exhibited a maximum activity at pH 7.5 in phosphate buffer, and adding flavin adenine dinucleotide to a final concentration of 15 μM would enhance the activity by three‐fold. Although this enzyme shares no more than 33% identity with any of reported 3‐hydroxybenzoate 6‐hydroxylases from Gram‐negative bacterial strains, there is little difference in subunit sizes and biochemical characteristics between them. (© 2010 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Molecular diversity of native rhizobia trapped by five field pea genotypes in Indian soils
Authors: Wadhwa K. , Dudeja S. S. , Yadav R. K. 
Published: August 2010

AbstractFive pea cultivars; HFP 4, HVP 3–5, HFP 9426, Jayanti and Hariyal, being grown in CCS Haryana Agricultural University farm were used to isolate native rhizobia. Selected 54 rhizobia, from all cultivars, were authenticated as rhizobia by plant infectivity test. Along with nodulation, symbiotic effectiveness in terms of symbiotic ratios showed wide range of effectiveness of pea rhizobia from 1.11 to 5.0. DNA of all the 54 rhizobia was extracted and amplified by PCR, using ERIC and 16S rDNA primers. Dendrogram based on ERIC profiles of these 54 rhizobia showed the formation of 13 subclusters at 80% level of similarity. Dendrogram based on RFLP of 16S rDNA by three restriction endonucleases; Msp I, Csp 6I and Rsa I; also formed 13 subclusters at 80% level of similarity. However, positioning of subclusters was different from that of ERIC based dendrogram. Majority of the isolates i.e. 64.8% by ERIC profiles and 44.4% by RFLP of 16S rDNA formed one cluster. Isolates from same nodule were not 100% similar. Considering each cluster representing a rhizobial genotype, both techniques used to assess molecular diversity indicated the presence of 13 genotypes of field pea rhizobia in CCS Haryana Agricultural University farm soil. Two pea rhizobial genotypes were able to nodulate all the five pea cultivars. Furthermore, high strain richness index (0.43–0.5) of field pea rhizobia was observed by both the techniques. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Validation of nested PCR and a selective biochemical method as alternatives for mycoplasma detection
Authors: Cheong Kyung Ah , Agrawal Santosh Rani , Lee Ai‐Young 
Published: August 2010

AbstractDirect culture is the most common way to reliably detect mycoplasma, but it is not practical for the qualitative control of cell therapeutics because of the elaborate culture medium, the prolonged incubation time, and the large sample volumes. Here, we chose two alternative methods using commercial detection kits, the PCR mycoplasma detection kit with nested PCR and the selective biochemical method, MycoAlert®, and validated them with the direct culture method as a reference. We tested eight mycoplasma species and five validation parameters: specificity, detection limit, robustness, repeatability, and ruggedness, based on the regulatory guidelines in the US Pharmacopoeia. All experiments were performed using fibroblasts spiked with mycoplasma. Specificity tests for both methods included all mycoplasma species, except Mycoplasma pneumonia and M. genitalium for the nested PCR and Ureaplasma urealyticum for the MycoAlert® assay. Regarding the detection limit, the nested PCR proved to be as sensitive as the direct culture method and more sensitive than the MycoAlert® assay. The predicted median for probit = 0.9 was 54 (44–76) CFU/ml for M. hyorhinis and 16 (13–23) CFU/ml for M. hominis by the nested PCR, but 431 (346–593) CFU/ml and 105 (87–142) CFU/ml, respectively, with MycoAlert®. Changes in the concentration of reagents, reagent lot, or individual analysts did not influence the results of the examined methods. The results of this study support nested PCR as a valuable alternative for mycoplasma detection. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)
Titanium particles activate toll‐like receptor 4 independently of lipid rafts in RAW264.7 murine macrophages
Authors: Islam Andrew S. , Beidelschies Michelle A. , Huml Anne , Greenfield Edward M. 
Published: August 2010

AbstractAdherent pathogen‐associated molecular patterns (PAMPs) act through toll‐like receptor 2 (TLR2) and TLR4 to increase the biological activity of orthopedic wear particles in cell culture and animal models of implant loosening. This study tested whether this is dependent on TLR association with lipid rafts as reported for the response to soluble TLR ligands. For this purpose, RAW264.7 murine macrophages were activated by exposure to titanium particles with adherent PAMPs, soluble lipopolysaccharide (LPS), soluble lipotecichoic acid (LTA), or heat‐killed bacteria that had been extensively washed to remove soluble PAMPs. Lipid rafts were isolated by two independent methods and the location of TLR4 and TLR2 was analyzed by Western blotting. The cognate TLRs associated with lipid rafts when the macrophages were activated with soluble LPS and LTA but not after stimulation with either titanium particles with adherent PAMPs or heat‐killed bacteria. The lipid raft disruptor, methyl‐β‐cyclodextrin, dose‐dependently inhibited TNF‐α release in response to LPS but had no affect on TNF‐α release in response to titanium particles with adherent PAMPs. We conclude, therefore, that titanium particles with adherent PAMPs and heat‐killed bacteria activate TLR2 and TLR4 in macrophages without inducing either TLR to associate with lipid rafts. These results have important implications for the mechanisms of orthopedic implant loosening as well the mechanisms for TLR activation in other inflammatory situations. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:211–217, 2011
Role of mitogen‐activated protein kinase in osteoblast differentiation
Authors: Lin Fu‐Hsiung , Chang Jessica B. , Brigman Brian E. 
Published: August 2010

AbstractLocal control of osteoblast differentiation and bone formation is not well understood. We have previously seen biphasic effects on cell differentiation in response to the short‐ and long‐term exposure to IL‐1β in rat calvarial osteoblasts. To characterize the signaling pathway mechanisms regulating IL‐1β biphasic effects, we examined the contribution of mitogen‐activated protein kinase (MAPK) family. Cells were pretreated with specific inhibitors to extracellular signal‐regulated kinase (ERK, PD98059), p38 (SB203580), and c‐JUN N‐terminal kinase (JNK, SP600125), then co‐cultured with IL‐1β for 2, 4, and 6 days. Cell differentiation was determined by measuring bone nodules after 10 days of culture. These inhibitors did not alter biphasic effects of IL‐1β on cell differentiation. However, PD98059 and U2016, another inhibitor of ERK activation robustly increased osteoblast differentiation compared to vehicle‐treated control in a time‐ and dose‐dependent manner. PD98059 appears to stimulate alkaline phosphatase (ALP) activity to promote cell differentiation, where IL‐1β appears to suppress it. Interestingly, continuous ERK inhibition with PD98059, after 2 and 4 days of IL‐1β treatment, enhanced the IL‐1β anabolic effect by increasing bone nodules formed. These observations provide a potential mechanism involving ERK pathway in osteoblasts differentiation and suggest that MAPK family may not directly regulate IL‐1β biphasic effects. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:204–210, 2011
Surgical synovectomy decreases density of sensory nerve fibers in synovial tissue of non‐inflamed controls and rheumatoid arthritis patients
Authors: Ossyssek B. , Anders S. , Grifka J. , Straub R.H. 
Published: August 2010

AbstractSurgical synovectomy is a technique to treat synovitis and pain in patients with rheumatoid arthritis (RA) resistant to DMARDs or therapy with biologics. Indication to synovectomy is subject to tight cooperation of orthopaedic surgeons and rheumatologists. It was thought that synovectomy leads to a reduction of sensory nerve fibers, called sensory denervation. Since sensory denervation after synovectomy has never been histologically tested, we aimed to investigate sensory and sympathetic innervation in synovial tissue before and after synovectomy. Eight non‐inflamed control subjects and eight patients with RA were included in this study with a two‐stage synovectomy approach (interval 40–50 days). Nerve fibers and cells in synovial tissue were detected and counted using immunofluorescence. Density of sympathetic nerve fibers did not change after synovectomy, whereas density of sensory nerve fibers decreased in all control subjects and seven of eight patients with RA. In parallel, the density of synovial cells increased after synovectomy in all control subjects and six of eight RA patients, which is indicative of a wound healing response. In one female RA patient, density of sensory nerve fibers increased and a very marked rise of cellular density was observed, too. This indicates that probably not all patients profit from surgical synovectomy. The majority of patients (94%) demonstrated sensory denervation after surgical synovectomy accompanied by a wound healing cell response. This study can help to explain the positive effects of surgical synovectomy which usually leads to pain reduction and improved mobility. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:297–302, 2011
Effects of botulinum toxin‐induced paralysis on postnatal development of the supraspinatus muscle
Authors: Das Rosalina , Rich Jason , Kim H. Mike , McAlinden Audrey , Thomopoulos Stavros 
Published: August 2010

AbstractThe mechanical environment plays an important role in musculoskeletal tissue development. The present study characterized changes in supraspinatus muscle due to removal of mechanical cues during postnatal development. An intramuscular injection of botulinum toxin type A (BTX) was used to induce and maintain paralysis in the left shoulders of mice since birth while the right shoulders received saline and served as contralateral controls. A separate group of animals was allowed to develop normally without any injections. Muscles were examined postnatally at various time points. The maximum isometric tetanic force generated by the muscle was significantly reduced in the BTX group compared to saline and normal groups. The paralyzed muscles were smaller and showed significant muscle atrophy and fat accumulation on histologic evaluation. Myogenic genes myogenin, myoD1, myf5, myf6, and fast type II myosin heavy chain (MHC) isoform were significantly upregulated while slow type I MHC isoform was significantly downregulated in the BTX group. Adipogenic genes C/EBPα, PPARγ2, leptin, and lipoprotein lipase were significantly upregulated in the BTX group. Results indicate that reduced muscle loading secondary to BTX‐induced paralysis leads to fat accumulation and muscle degeneration in the developing muscle. Understanding the molecular and compositional changes in developing supraspinatus muscles may be useful for identifying and addressing the pathological changes that occur in shoulder injuries such as neonatal brachial plexus palsy. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:281–288, 2011
Background removal from polarized Raman spectra of tooth enamel using the wavelet transform
Authors: Li Jianghua , Choo‐Smith Lin‐P'ing , Tang Zhilie , Sowa Michael G. 
Published: August 2010

AbstractA wavelet transformation method is introduced to remove the large fluorescence background from polarized Raman spectra of stained tooth enamel. This method exploits the wavelet multiresolution decomposition where the experimental Raman spectrum is decomposed into signals with different frequency components, and where the lowest frequency background and highest frequency noise are removed. This method is optimized using a simulated collection of parallel‐polarized and cross‐polarized Raman spectra of the enamel and then applied to a set of experimental data. The results show that the wavelet transform technique can extract the pure spectra from background and noise, with the depolarization ratio used to discriminate between early dental caries and sound enamel preserved. Copyright © 2010 Crown in the right of Canada. Published by John Wiley & Sons, Ltd.
Using Raman spectroscopy to monitor surface finish and roughness of components manufactured by selective laser sintering
Authors: Beard M. A. , Ghita O. R. , Evans K. E. 
Published: August 2010

AbstractSelective laser sintering (SLS) is an additive manufacturing process used to realise fully functional component manufacture. Numerous parameters are used in the process to control variables such as laser power, scan speed, laser spot size and overlap of scan vectors. All of these parameters can dramatically alter the sintering process and therefore final component properties. This paper presents how Raman spectroscopy intensity effects, caused by the surface roughness of the components produced, can be used to monitor the degree of sintering between particles in the SLS process. Copyright © 2010 John Wiley & Sons, Ltd.
Visualization of β‐carotene and starch granules in plant cells using CARS and SHG microscopy
Authors: Brackmann Christian , Bengtsson Anton , Alminger Marie Larsson , Svanberg Ulf , Enejder Annika 
Published: August 2010

AbstractInformation on the content and bioavailability of provitamin A carotenoids, such as β‐carotene, in plant foods is of great interest due to the widespread vitamin A deficiency in developing countries. While the amount of β‐carotene can readily be quantified with analytical techniques, there is limited information on β‐carotene morphology in native plant materials. Here, we introduce nonlinear microscopy for three‐dimensional, label‐free imaging of carotenoids in fresh and thermally treated plant tissues, providing quantitative information at single‐aggregate level and detailed insight into their distribution. Carotenoids in orange‐fleshed sweet potato (OFSP), carrot, and mango were visualized by coherent anti‐Stokes Raman scattering (CARS) microscopy and, in the case of OFSP, related to the plant‐matrix morphology by simultaneous second‐harmonic generation (SHG) microscopy of starch granules. Sizes, shapes, densities, and location of different types of carotenoid bodies were quantified. While OFSP and carrot showed heterogeneous rod‐shaped bodies with high carotenoid densities indicated by higher CARS signals, the carotenoid‐filled lipid droplets in mango appeared as homogeneous low‐density aggregates of rounded shape. In addition, β‐carotene densities and morphologies in OFSP were studied after thermal processing, showing that the bodies remain intact despite significant changes of the surrounding starch granules. Copyright © 2010 John Wiley & Sons, Ltd.
Surface‐enhanced Raman spectroscopy of DNA from leaves of in vitro grown apple plants
Authors: Muntean Cristina M. , Leopold Nicolae , Halmagyi Adela , Valimareanu Sergiu 
Published: August 2010

AbstractUltrasensitive Raman measurements of nucleic acids are possible by exploiting the effect of surface‐enhanced Raman scattering (SERS). In this work, the vibrational spectra of eight genomic DNAs from in vitro grown apple leaf tissues (Malus domestica Borkh., Fam Rosaceae, cvs. Florina, Idared, Rebra, Goldrush, Romus 3, Romus 4 and the rootstocks M9 and M26) were analyzed using surface‐enhanced Raman spectroscopy, in the wavenumber range 200–1800 cm−1. SERS signatures, spectroscopic band assignments and structural interpretations of these plant genomic DNAs are reported. Strong dependences of the SERS spectra on genomic DNA amount in the measured sample volume and on time were observed. Similarities of the SERS signals of DNAs from Rebra and Romus 3 leaves were detected. To our knowledge, this is the first SERS study on genomic DNA from leaf tissues. The present work provides a basis for future use of surface‐enhanced Raman spectroscopy to analyze specific plant DNA–ligand interactions or DNA structural changes induced by plants' stress conditions associated with their natural environment. Besides, this study will generate information that is valuable in the development of low‐level plant DNA‐based analytical sensors. Copyright © 2010 John Wiley & Sons, Ltd.
New HPLC‐chemometric approaches to the analysis of isoflavones in Trifolium lucanicum Gasp.
Authors: Küçükboyacı Nurgün , Güvenç Ayşegül , Dinç Erdal , Adıgüzel Nezaket , Bani Barış 
Published: August 2010

AbstractNew HPLC‐chemometric approaches were proposed for the simultaneous chromatographic quantification of daidzein, genistein, formononetin, and biochanin A in the samples consisting of the aerial parts of Trifolium lucanicum Gasp. (Leguminosae). Partial least squares and principal component regression algorithms were applied to the multiple chromatographic data set obtained by measuring at 240, 248, 256, and 264 nm to construct HPLC‐partial least squares and HPLC‐principal component regression calibrations. Chromatographic separation was carried out by using a mobile phase containing methanol, acetate buffer (pH=4.75) and acetonitrile (21:58:21, v/v/v) on the reversed phase column, Supelcosil™ LC‐18 (15 cm×4.6 mm id). In addition, conventional HPLC based on the detection at a single wavelength was used for the determination of each compound in the extracts of T. lucanicum. The validity and applicability of the proposed HPLC‐chemometric and conventional HPLC methods were performed by analyzing various synthetic plant samples. A good agreement was observed in the application of the proposed HPLC‐chemometric tools to the synthetic and extracted samples of T. lucanicum.
Analysis of Aloe‐based phytotherapeutic products by using nano‐LC‐MS
Authors: Fanali Salvatore , Aturki Zeineb , D'Orazio Giovanni , Rocco Anna , Ferranti Anna , Mercolini Laura , Raggi Maria Augusta 
Published: August 2010

AbstractThis article proposes a chromatographic method for the analysis of extracts of Aloe plants. The method was developed with a laboratory assembled nano‐LC system coupled with a UV detector, followed by an IT‐mass spectrometer. With a step gradient mode of ACN/H2O mixtures and employing a capillary column packed with C18 (100 μm id), a complete separation of the following anthrones was achieved: aloin (in its two isomeric forms A and B), 5‐hydroxyaloin and 7‐hydroxyaloin (in its two isomeric forms A and B). The optimized nano‐LC‐MS method was validated for the quantification of aloin, the main component of Aloe with known pharmacological activities. RSD values obtained for retention time and peak areas were 1.3 and 12.1%, respectively. LOD and LOQ values of 0.4 and 1.5 μg/mL were obtained for each aloin isomer. The method was applied to the analysis of Aloe vera and A. ferox extracts in order to acquire a fingerprint, characteristic for each plant. Several phenolic compounds were detected by UV and identified by MS. A. vera and A. ferox showed different profiles and it was possible to discriminate them. Several commercial formulations, declared to contain Aloe extracts, were analyzed. Comparing their chromatograms with those obtained from A. vera and A. ferox, it was possible to recognize the Aloe species and to determine aloin.