Identification of genes induced by salt stress from Medicago truncatula L. seedlings

In order to identify genes induced during the salt stress response in barrel medic (Medicago truncatula L) seedlings, a cDNA library by salt stress was constructed  by suppression subtractive hybridization (SSH). Total RNA from 15-day-old seedlings was used as a ‘driver’, and total RNA from seedlings induced by salt was used as a ‘tester’. One hundred and sixty nine clones identified as positive clones by reverse northern dot-blotting resulted in 75 uni-ESTs that comprised of 13 contigs  and 62 singletons. Basic Local Alignment Search Tool (BLAST) analysis of deduced protein sequences revealed that 35 expressed sequence tags (ESTs) had identity similar to proteins with known function, while 27 could not be annotated at all. Most of the known function sequences were homologous to genes involved in abiotic stress in plants. Among these protein, citrate synthase, ribulose- 1,5-bisphosphate carboxylase, chloroplast protein, phosphoenolpyruvate carboxylase and  chloroplast outer envelope protein are related to photosynthesis; DNA binding/transcription factor, putative AP2/EREBP transcription factor, Cab9 gene, photosystem II polypeptide and calcium-dependent protein kinase play a significant role in signal transduction and transcription regulation; and aldolase and sucrose synthase are interrelated to osmolyte synthesis. Moreover, 5 of the ESTs, similar to genes from other plant species and closely involved in salt stress were isolated from M. truncatula L. They are superoxide dimutase (SOD)-1, gene for copper/zinc superoxide dismutase, cysteine protease, Na+/H+ antiporter and salt overly sensitive 2 (SOS2). To further assess the expression level of salt-induced ESTs, real-time polymerase chain reaction (PCR) analysis was employed, and the result showed that these genes have significantly increased expression and probably play an important role in the response of plants to salt stress.Key words: Barrel medic (Medicago truncatula L.), suppression subtraction hybridization (SSH), reverse northern dot-blotting, salt stress, real-time polymerase chain reaction (PCR)

Spatial Discretization for Stochastic Semi-Linear Subdiffusion Equations Driven by Fractionally Integrated Multiplicative Space-Time White Noise

From MDPI via Jisc Publications RouterHistory: accepted 2021-08-03, pub-electronic 2021-08-12Publication status: PublishedSpatial discretization of the stochastic semi-linear subdiffusion equations driven by fractionally integrated multiplicative space-time white noise is considered. The nonlinear terms f and σ satisfy the global Lipschitz conditions and the linear growth conditions. The space derivative and the fractionally integrated multiplicative space-time white noise are discretized by using the finite difference methods. Based on the approximations of the Green functions expressed by the Mittag–Leffler functions, the optimal spatial convergence rates of the proposed numerical method are proved uniformly in space under some suitable smoothness assumptions of the initial value

Rapid determination of 103 common veterinary drug residues in milk and dairy products by ultra performance liquid chromatography tandem mass spectrometry

A multi-residue method has been developed for the identification and quantification of 103 common veterinary drug residues in milk and dairy Products. This method was based on QuEChERS with dispersive solid-phase where C18 sorbent and anhydrous sodium sulfate were used to sample purification. After evaporation and reconstitution, the samples were analyzed by ultra-performance liquid chromatography-tandem mass spectrometry. The mean recovery results were all higher than 60% except ampicillin, pipemidic acid, enoxacin, and estriol, and the relative standard deviation was <20.0%. The limit of quantification ranged between 0.1 and 5 μg/kg for milk and between 0.5 and 25 μg/kg for milk powder. It was successfully used to detect residues of veterinary drug in real samples. This study proposes a simple and fast analytical method for monitoring multi-class veterinary drug residues to ensure food safety

Detection of Pine Nut Allergen in Three Kinds of Food by Ultra-high Performance Liquid Chromatography-Tandem Mass Spectrometry

An ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was established for the detection of pine nut allergen Pin k 2 in food matrices. Pine nuts were ground, degreased, and enzymatically extracted and the hydrolysate was separated and analyzed by using an Easy-nLC 1000-QExecutive high-resolution mass spectrometer, and the mass spectral data obtained were processed using Protein Pilot TM software and the Uniprot protein database. The specificity was verified by Basic Local Alignment Search Tool (BLAST), and three pine nut-specific peptides were selected finally. The developed method exhibited a good linear relationship in the concentration range of 0.001–50 mg/mL, and the limit of quantification was 1 mg/kg. The average recoveries for blank biscuit, chocolate and beverage were 88.50%–107.57%, with a relative standard deviation (RSD) not exceeding 6.08%, and the matrix effect was 89.77%–96.13%. This method has the advantages of high sensitivity and good specificity, and can be applied to the detection of pine nut allergens in food samples such as biscuits, chocolate, and beverages, which provides technical support for the authentication of food labels and the detection of latent allergens in foods

Biochemical characterization of an acetylesterase from Bacillus subtilis and its application for 7-aminocephalosporanic acid deacetylation

Deacetyl-7-aminocephalosporanic acid (D-7-ACA), which could be converted from 7-aminocephalosporanic acid (7-ACA), is a crucial starting material that is used for synthesizing industrial semisynthetic β-lactam antibiotics. Enzymes involved in the conversion from 7-ACA to D-7-ACA present critical resources in the pharmaceutical industry. In the present study, a putative acetylesterase, EstSJ, identified from Bacillus subtilis KATMIRA1933, was first heterologously expressed in Escherichia coli BL21(DE3) cells and biochemically characterized. EstSJ belongs to carbohydrate esterase family 12 and is active on short-chain acyl esters from p-NPC2 to p-NPC6. Multiple sequence alignments showed that EstSJ was also an SGNH family esterase with a typical GDS(X) motif at its N-terminal end and a catalytic triad composed of Ser186-Asp354-His357. The purified EstSJ displayed the highest specific activity of 1,783.52 U mg–1 at 30°C and pH 8.0, and was stable within the pH range of 5.0–11.0. EstSJ can deacetylate the C3′ acetyl group of 7-ACA to generate D-7-ACA, and the deacetylation activity was 4.50 U mg–1. Based on the structural and molecular docking with 7-ACA, the catalytic active sites (Ser186-Asp354-His357) together with four substrate-binding residues (Asn259, Arg295, Thr355, and Leu356) of EstSJ are revealed. This study provided a promising 7-ACA deacetylase candidate that could be applied to produce D-7-ACA from 7-ACA in the pharmaceutical industry

Evaluation of a Novel Biphasic Culture Medium for Recovery of Mycobacteria: A Multi-Center Study

on L-J slants. Automated liquid culture systems are expensive. A low-cost culturing medium capable of rapidly indicating the presence of mycobacteria is needed. The aim of this study was to develop and evaluate a novel biphasic culture medium for the recovery of mycobacteria from clinical sputum specimens from suspected pulmonary tuberculosis patients.<0.001).

Co-Deletion of Chromosome 1p/19q and IDH1/2 Mutation in Glioma Subsets of Brain Tumors in Chinese Patients

OBJECTIVE: To characterize co-deletion of chromosome 1p/19q and IDH1/2 mutation in Chinese brain tumor patients and to assess their associations with clinical features. METHODS: In a series of 528 patients with gliomas, pathological and radiological materials were reviewed. Pathological constituents of tumor subsets, incidences of 1p/19q co-deletion and IDH1/2 mutation in gliomas by regions and sides in the brain were analyzed. RESULTS: Overall, 1p and 19q was detected in 339 patients by FISH method while the sequence of IDH1/2 was determined in 280 patients. Gliomas of frontal, temporal and insular origin had significantly different pathological constituents of tumor subsets (P<0.001). Gliomas of frontal origin had significantly higher incidence of 1p/19q co-deletion (50.4%) and IDH1/2 mutation (73.5%) than those of non-frontal origin (27.0% and 48.5%, respectively) (P<0.001), while gliomas of temporal origin had significantly lower incidence of 1p/19q co-deletion (23.9%) and IDH1/2 mutation (41.7%) than those of non-temporal origin (39.9% and 63.2%, respectively) (P = 0.013 and P = 0.003, respectively). Subgroup analysis confirmed these findings in oligoastrocytic and oligodendroglial tumors, respectively. Although the difference of 1p/19q co-deletion was not statistically significant in temporal oligodendroglial tumors, the trend was marginally significant (P = 0.082). However, gliomas from different sides of the brain did not show significant different pathological constituents, incidences of 1p/19q co-deletion or IDH1/2 mutation. CONCLUSION: Preferential distribution of pathological subsets, 1p/19q co-deletion and IDH1/2 mutation were confirmed in some brain regions in Chinese glioma patients, implying their distinctive tumor genesis and predictive value for prognosis

Isolation and functional characterization of a Medicago sativa L. gene, MsLEA3-1

A full-length cDNA of 1,728 nt, called MsLEA3-1, was cloned from alfalfa by rapid amplification of cDNA ends from an expressed sequence tag homologous to soybean pGmPM10 (accession No. AAA91965.1). MsLEA3-1, encodes a deduced protein of 436 amino acids, a calculated molecular weight of 47.0 kDa, a theoretical isoelectric point of 5.18, and closest homology with late embryogenesis abundant proteins in soybean. Sequence homology suggested a signal peptide in the N terminus, and subcellular localization with GFP revealed that MsLEA3-1 was localized preferentially to the nucleolus. The transcript titre of MsLEA3-1 was strongly enriched in leaves compared with roots and stems of mature alfalfa plants. Gene expression of MsLEA3-1 was strongly induced when seedlings were treated with NaCl and ABA. Expression of the MsLEA3-1 transgenic was detected in transgenic tobacco. Malondialdehyde content and, electrical conductivity content were reduced and electrical conductivity and proline content were increased in transgenic tobacco compared with non-transgenic tobacco under salt stress. The results showed that accumulation of the MsLEA3-1 protein in the vegetative tissues of transgenic plants enhanced their tolerance to salt stress. These results demonstrate a role for the MsLEA3-1 protein in stress protection and suggest the potential of the MsLEA3-1 gene for genetic engineering of salt tolerance