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#30240699   2018/09/18 To Up

Identification and sequence analysis of two novel cryptic plasmids isolated from the vaginal mucosa of South African women.

The vaginal mucosa is dominated by Gram positive, rod shaped lactobacilli which serve as a natural barrier against infection. In both healthy- and bacterial vaginosis (BV)-infected women Lactobacillus crispatus and Lactobacillus jensenii have been found to be the predominant Lactobacillus species. Many studies have been conducted to assess factors influencing lactobacilli dominance in the vaginal microbiome. In the present study two plasmids, pLc4 and pLc17, isolated from vaginal Lactobacillus strains of both healthy and BV-infected women were characterized. The smaller plasmid, pLc4 (4224 bp), was detected in both L. crispatus and L. jensenii strains, while pLc17 was only detected in L. crispatus. Based on its nucleotide sequence pLc4 appears highly novel, with its replication protein having 44% identity to the replication initiation protein of pSMQ173b_03. Phylogenetic analysis with other Rolling Circle Replication plasmids confirmed that pLc4 shows a low degree of similarity to these plasmids. Plasmid pLc17 (16,663 bp) appears to carry both a RCR replicon and a theta replicon, which is rare in naturally occurring plasmids. pLc4 was maintained at a high copy number of 29, while pLc17 appears to be a medium copy number plasmid maintained at 11 copies per chromosome. While sequence analysis is a valuable tool to study cryptic plasmids, further function-based analysis will be required in order to fully elucidate the role of these plasmids within the vaginal milieu.
Lyle Harris, Leonardo J van Zyl, Bronwyn M Kirby-McCullough, Leonard H Damelin, Caroline T Tiemessen, Marla Trindade

1355 related Products with: Identification and sequence analysis of two novel cryptic plasmids isolated from the vaginal mucosa of South African women.

5 G1 module1 module25 mg 15 ml 500 Units1 module10 mg100ug

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#30175516   2018/10/11 To Up

Phosphatidylinositol-hydrolyzing phospholipase C4 modulates rice response to salt and drought.

Phosphatidylinositol-specific phospholipase C (PI-PLC) is involved in stress signalling but its signalling function remains largely unknown in crop plants. Here, we report that the PI-PLC4 from rice (Oryza sativa cv), OsPLC4, plays a positive role in osmotic stress response. Two independent knockout mutants, plc4-1 and plc4-2, exhibited decreased seedling growth and survival rate whereas overexpression of OsPLC4 improved survival rate under high salinity and water deficiency, compared with wild type (WT). OsPLC4 hydrolyses PI, phosphatidylinositol 4-phosphate (PI4P), and phosphatidylinositol-4,5-bisphosphate (PIP ) to generate diacylglycerol (DAG) in vitro. Knockout of OsPLC4 attenuated salt-induced increase of phosphatidic acid (PA) whereas overexpression of OsPLC4 decreased the level of PI4P and PIP under salt treatment. Applications of DAG or PA restored the growth defect of plc4-1 to WT but DAG kinase inhibitor 1 blocked the complementary effect of DAG in plc4-1 under salt stress. In addition, the loss of OsPLC4 compromised the increase of inositol triphosphate and free cytoplasmic Ca ([Ca ] ) and inhibited the induction of genes involved in Ca sensor and osmotic stress response to salt stress. The results indicate that OsPLC4 modulates the activity of two signalling pathways, PA and Ca , to affect rice seedling response to osmotic stress.
Xianjun Deng, Shu Yuan, Huasheng Cao, Sin Man Lam, Guanghou Shui, Yueyun Hong, Xuemin Wang

2826 related Products with: Phosphatidylinositol-hydrolyzing phospholipase C4 modulates rice response to salt and drought.

1.0 g2.5 g250 mg5 mg 500 G1 mg2.5 mg1 mg 1 G1 g100 mg1 mg

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#28102910   2017/03/20 To Up

Arabidopsis phosphoinositide-specific phospholipase C 4 negatively regulates seedling salt tolerance.

Previous physiological and pharmacological studies have suggested that the activity of phosphoinositide-specific phospholipase C (PI-PLC) plays an important role in regulating plant salt stress responses by altering the intracellular Ca concentration. However, the individual members of plant PLCs involved in this process need to be identified. Here, the function of AtPLC4 in the salt stress response of Arabidopsis seedlings was analysed. plc4 mutant seedlings showed hyposensitivity to salt stress compared with Col-0 wild-type seedlings, and the salt hyposensitive phenotype could be complemented by the expression of native promoter-controlled AtPLC4. Transgenic seedlings with AtPLC4 overexpression (AtPLC4 OE) exhibited a salt-hypersensitive phenotype, while transgenic seedlings with its inactive mutant expression (AtPLC4m OE) did not exhibit this phenotype. Using aequorin as a Ca indicator in plc4 mutant and AtPLC4 OE seedlings, AtPLC4 was shown to positively regulate the salt-induced Ca increase. The salt-hypersensitive phenotype of AtPLC4 OE seedlings was partially rescued by EGTA. An analysis of salt-responsive genes revealed that the transcription of RD29B, MYB15 and ZAT10 was inversely regulated in plc4 mutant and AtPLC4 OE seedlings. Our findings suggest that AtPLC4 negatively regulates the salt tolerance of Arabidopsis seedlings, and Ca may be involved in regulating this process.
Keke Xia, Bo Wang, Jiewei Zhang, Yuan Li, Hailian Yang, Dongtao Ren

2398 related Products with: Arabidopsis phosphoinositide-specific phospholipase C 4 negatively regulates seedling salt tolerance.

2.5 g1 g1 g100 mg1 g 5 G5 g 25 G1 g0.2 mg25 mg25 mg

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#22899083   2012/08/17 To Up

Defense activation triggers differential expression of phospholipase-C (PLC) genes and elevated temperature induces phosphatidic acid (PA) accumulation in tomato.

Recently, we provided the first genetic evidence for the requirement of tomato PLC4 and PLC6 genes in defense activation and disease resistance. The encoded enzymes were catalytically active as they were able to degrade phosphatidylinositol (PI), thereby producing diacylglycerol (DG). Here we report differential PLC gene expression following the initiation of defense signaling by the interaction between Cladosporium fulvum resistance (R) protein Cf-4 and its matching effector Avr4 in tomato hybrid seedlings that express both Cf-4 and Avr4. Furthermore, we observed that PLC3 and PLC6 gene expression is upregulated by elevated temperature in the control seedlings. This upregulation coincides with an increase in the levels of phosphatidic acid (PA) and a decrease in the levels of PI and phosphatidylinositol phosphate (PIP). The decrease in PI and PIP levels matches with the activation of PLC. In addition, the levels of the structural phospholipids phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylglycerol (PG) declined transiently during recovery after the exposure to elevated temperature., Further studies will be required to explain the mechanism causing the sustained accumulation of PA during recovery, combined with a reduction in the levels of structural phospholipids.
Ahmed Abd-El-Haliem, Harold J G Meijer, Wladimir I L Tameling, Jack H Vossen, Matthieu H A J Joosten

1791 related Products with: Defense activation triggers differential expression of phospholipase-C (PLC) genes and elevated temperature induces phosphatidic acid (PA) accumulation in tomato.

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#11443096   // To Up

Regulation of Staphylococcus aureus type 5 and type 8 capsular polysaccharides by CO(2).

Staphylococcus aureus expression of capsular polysaccharide type 5 (CP5) has been shown to be downregulated by CO(2). Here we show that CO(2) reduces CP5 expression at the transcriptional level and that CO(2) regulates CP8 expression depending on the genetic background of the strains. Growth in the presence of air supplemented with 5% CO(2) caused a significant decrease in CP8 expression in four S. aureus strains, a marginal effect in four strains, and higher CP8 expression in strain Becker. Absolute CP8 expression in the nine S. aureus strains differed largely from strain to strain. Four groups of strains were established due to sequence variations in the promoter region of cap5 and cap8. To test whether these sequence variations are responsible for the different responses to CO(2), promoter regions from selected strains were fused to the reporter gene xylE in pLC4, and the plasmids were electrotransformed into strains Becker and Newman. XylE activity was negatively regulated by CO(2) in all derivatives of strain Newman and was always positively regulated by CO(2) in all derivatives of strain Becker. Differences in promoter sequences did not influence the pattern of CP8 expression. Therefore, the genetic background of the strains rather than differences in the promoter sequence determines the CO(2) response. trans-acting regulatory molecules may be differentially expressed in strain Becker versus strain Newman. The strain dependency of the CP8 expression established in vitro was also seen in lung tissue sections of patients with cystic fibrosis infected with CP8-positive S. aureus strains.
S Herbert, S W Newell, C Lee, K P Wieland, B Dassy, J M Fournier, C Wolz, G Döring

2257 related Products with: Regulation of Staphylococcus aureus type 5 and type 8 capsular polysaccharides by CO(2).

96T50ul50ug 50μl50μl500 100ul1 ml

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#2045363   // To Up

The Escherichia coli hemL gene encodes glutamate 1-semialdehyde aminotransferase.

delta-Aminolevulinic acid (ALA), the first committed precursor of porphyrin biosynthesis, is formed in Escherichia coli by the C5 pathway in a three-step, tRNA-dependent transformation from glutamate. The first two enzymes of this pathway, glutamyl-tRNA synthetase and Glu-tRNA reductase, are known in E. coli (J. Lapointe and D. Söll, J. Biol. Chem. 247:4966-4974, 1972; D. Jahn, U. Michelsen, and D. Söll, J. Biol. Chem. 266:2542-2548, 1991). Here we present the mapping and cloning of the gene for the third enzyme, glutamate 1-semialdehyde (GSA) aminotransferase, and an initial characterization of the purified enzyme. Ethylmethane sulfonate-induced mutants of E. coli AB354 which required ALA for growth were isolated by selection for respiration-defective strains resistant to the aminoglycoside antibiotic kanamycin. Two mutations were mapped to min 4 at a locus named hemL. Map positions and resulting phenotypes suggest that hemL may be identical with the earlier described porphyrin biosynthesis mutation popC. Complementation of the auxotrophic phenotype by wild-type DNA from the corresponding clone pLC4-43 of the Clarke-Carbon bank (L. Clarke and J. Carbon, Cell 9:91-99, 1976) allowed the isolation of the gene. Physical mapping showed that hemL mapped clockwise next to fhuB. The hemL gene product was overexpressed and purified to apparent homogeneity. The pure protein efficiently converted GSA to ALA. The reaction was stimulated by the addition of pyridoxal 5' -phosphate or pyridoxamine 5' -phosphate and inhibited by gabaculine or aminooxyacetic acid. The molecular mass of the purified GSA aminotransferase under denaturing conditions was 40,000 Da, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme has apparent native molecular mass of approximately 80,000 Da, as determined by rate zonal sedimentation on glycerol gradients and molecular sieving through Superose 12, which indicates a homodimeric alpha2, structure of the protein.
L L Ilag, D Jahn, G Eggertsson, D Söll

2496 related Products with: The Escherichia coli hemL gene encodes glutamate 1-semialdehyde aminotransferase.

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#2001999   // To Up

Partial characterization of a lysU mutant of Escherichia coli K-12.

The Escherichia coli K-12 strain GNB10181 shows no inducible lysyl-tRNA synthetase (LysRS) activity. Two-dimensional gel electrophoretic analysis of the polypeptides synthesized by this strain indicates that the normal lysU gene product, LysU, is absent. When both GNB10181 and its parent, MC4100, were grown at elevated temperatures (42 to 45 degrees C) no significant difference between their growth rates was observed. The lysU mutation was transferred to other E. coli K-12 backgrounds by using P1 transduction. The lysU transductants behaved comparably to their lysU+ parents at different growth temperatures. Therefore, the LysU proteins does not appear to be essential for growth at high temperatures, at least under the conditions examined here. In addition, lysU transductants were found to be defective for inducible lysine decarboxylase, (LDC), inducible arginine decarboxylase (ADI), and melibiose utilization (Mel), which are all missing in GNB10181. Complementation of the above missing functions was achieved by using the Clarke-Carbon plasmids pLC4-5 (LysU LDC) and pLC17-38 (LysU Mel ADI). From these experiments, it appears that GNB10181 has suffered a chromosomal deletion between 93.4 and 93.7 min, which includes the lysU gene. By using plasmid pLC17-38, the position of ADI on two-dimensional gels was identified. Finally, lysS delta lysU double mutants were constructed which can potentially be used as positive selection agents for the isolation of LysRS genes from other sources.
M Hassani, M V Saluta, G N Bennett, I N Hirshfield

1987 related Products with: Partial characterization of a lysU mutant of Escherichia coli K-12.

200 200 200 200 200 100 100 200 1 mL200 200 1 mL

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#2162159   // To Up

Molecular cloning of feline leukemia provirus genomes integrated in the feline large granular lymphoma cells.

Feline leukemia virus (FeLV) is a horizontally transmitted agent of the domestic cat which is known to be associated with wide spectrum of diseases of the hematopoietic system. In the present study, proviral DNAs of FeLV proviruses were examined in the tumor cells of natural killer cell lineage which is very rare in cats. In the chromosomal DNA of the tumor cells, 5 distinct bands corresponding to exogenous FeLV provirus genomes were detected by digestion with EcoRI which does not cut most FeLV isolates. Five clones of pLC1, pLC2, pLC3, pLC4, and pLC5 obtained from the 5 respective bands were analysed by restriction endonuclease mapping and Southern blot hybridization using gene-specific probes of FeLV. The results have clearly demonstrated that pLC4 and pLC5 contained large deletions in the pol and part of gag regions, while the full-length proviruses could be observed in pLC1 and pLC2. Furthermore, pLC3 contained part of a variant FeLV genome having an EcoRI site in its gag region. The molecular clones of defective and variant FeLV in this study may be useful for the further examination of tumorigenesis of large granular lymphoma in the cat.
Y Matsumoto, H Tsujimoto, M Fukasawa, Y Hirota, T Miura, M Hayami, R Goitsuka, K Ono, A Hasegawa

1459 related Products with: Molecular cloning of feline leukemia provirus genomes integrated in the feline large granular lymphoma cells.

100 1 mL250 ug100 10 rxns1 mL250 ug1 mL1.00 flask250 ug1 mL250 ug

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