ABSTRACT: nano-DESI analysis of S. griseoflavus colonies on agar medium. S. griseoflavus WT, a hormaomycin overproducing and a hormaomycin deficient mutant were analyzed.
Project description:Total protein and membrane-enriched fractions of Synechocystis sp. PCC 6803 wild type (WT) and mutant strains affected with inactivated DNA methyltransferase genes were analyzed by LC-HDMSE. The mutation of the DNA methyltransferase gene sll0729 encoding M.Ssp6803II initially led to a strong phenotype, including a lowered chlorophyll/phycocyanin ratio, impaired growth, and alterations in gene expression. Prolonged cultivation revealed instability of the initially obtained phenotype. Colonies showing normal pigmentation and WT-like growth appeared regularly and in high frequencies on agar plates. These colonies represent suppressor mutants, since the sll0729 gene is still completely inactivated and methylation of the M.Ssp6803II target GGCC sites does not occur. The proteomic analysis comprises the WT, two strains of suppressor mutants sll0729_1 and sll0729_15 as well as mutants of the DNA methyltransferase genes slr6095 and sll8009. For each of these five strains we investigated three biological replicates. Comparisons between the WT and these two suppressor mutant strains, but also between them and the slr6095 and sll8009 mutants enabled the detection of expression differences, which are specifically linked to the absence of M.Ssp6803II-related DNA methylation.
Project description:Total protein and membrane-enriched fractions of Synechocystis sp. PCC 6803 wild type (WT) and mutant strains affected with inactivated DNA methyltransferase genes were analyzed by LC-HDMSE. The mutation of the DNA methyltransferase gene sll0729 encoding M.Ssp6803II initially led to a strong phenotype, including a lowered chlorophyll/phycocyanin ratio, impaired growth, and alterations in gene expression. Prolonged cultivation revealed instability of the initially obtained phenotype. Colonies showing normal pigmentation and WT-like growth appeared regularly and in high frequencies on agar plates. These colonies represent suppressor mutants, since the sll0729 gene is still completely inactivated and methylation of the M.Ssp6803II target GGCC sites does not occur. The proteomic analysis comprises the WT, two strains of suppressor mutants sll0729_1 and sll0729_15 as well as mutants of the DNA methyltransferase genes slr6095 and sll8009. For each of these five strains we investigated three biological replicates. Comparisons between the WT and these two suppressor mutant strains, but also between them and the slr6095 and sll8009 mutants enabled the detection of expression differences, which are specifically linked to the absence of M.Ssp6803II-related DNA methylation.
Project description:12plex_medicago_2013-08 - r108 in symbiosis with rhizobia wt or rhizobia mutant for baca. - Two experiments to compare the transcriptomic response of medicago plants: Agar medium versus Phytagel medium (exp1) and rhizobium WT versus BacA (exp2). - Medicago truncatula ecotype R108 was inoculated with the symbiotic rhizobium Sinorhizobium meliloti strain Sm1021 and with its derivative mutant delta bacA. Nodules were collected 13 days after inoculation, and RNA were prepared for transcriptome analysis, there were three biological independant experiements.
Project description:nanoDESI-MS analysis of Streptomyces coelicolor colonies on agar medium. The effect of an actinobacterial signaling metabolite on Streptomyces coelicolor was tested.
Project description:nanoDESI-MS analysis of Streptomyces griseus colonies on agar medium. The effect of an actinobacterial signaling metabolite on Streptomyces griseus was tested.
Project description:nanoDESI-MS analysis of Streptomyces roseosporus colonies on agar medium. The effect of an actinobacterial signaling metabolite on Streptomyces roseosporus was tested.
Project description:12plex_medicago_2013-08 - r108 permissive medium versus non permissive medium. - Two experiments to compare the transcriptomic response of medicago plants: Agar medium versus Phytagel medium (exp1) and rhizobium WT versus BacA (exp2). - Medicago truncatula R108 seedlings were inoculated with S. medicae WSM419 and were cultivated during three days on buffered nidulation medium solidified with Phytagel or Agar.
Project description:The response regulator MtrA is highly conserved in Streptomyces species where it coordinates antibiotic production with sporulation. Loss of MtrA results in a conditional bald phenotype in S. avermitilis, S. coelicolor and S. venezuelae, where the production of aerial hyphae and spores is blocked on some but not all growth media. In S. venezuelae NRRL-B-65442 ChIP-seq showed that MtrA binds to the promoters of >1000 genes including key genes required for development. Here we show that an S. venezuelae NRRL-B-65442 ∆mtrA mutant is bald on R2YE agar but develops normally on MYM agar. We used an in vitro DNA footprinting technique called Reusable DNA Capture Technology combined with Surface Plasmon Resonance (ReDCaT SPR) to identify the precise MtrA binding sites at the promoters of the developmental genes adpA, bldM, dnaA, filP, mtrA, ssgB and whiI. The results show that purified MtrA binds to single sites upstream of filP, mtrA, ssgB and whiI and to multiple sites upstream of adpA (x2), bldM (x5) and dnaA (x3) which may be indicative of the complex regulation of these genes. Tandem mass tag proteomics on the wild-type and ∆mtrA strains revealed that the levels of BldM and WhiI are 4-fold and 7-fold lower in the ∆mtrA mutant grown on R2YE agar suggesting they are directly regulated by MtrA under these growth conditions. They were not significantly affected in cultures grown on MYM agar. The expression of the key BldM and WhiI target genes ssgB, ssgR and whiB, which are key to development is also significantly reduced on R2YE agar. We conclude that MtrA directly activates expression of bldM and whiI on R2YE agar and that reduction in the levels of these regulators leads to the bald phenotype of the ∆mtrA mutant on this growth medium.
Project description:au07-03_myb77 - wt/myb77 - Transcriptome analysis of Myb77 knock out mutant line or of Myb77fragment overxpressing transgenic line. - Plants were grown in vitro on 1/2 MS medium, 3% sucrose, 0.8% agar for 21 days under balance day cycle (12 hours daylight at 22°C; 12 hours dark at 20°C) in climate chamber. The whole plants were collected for RNAs extraction. Keywords: gene knock in (transgenic),gene knock out
Project description:In this experiment the transcriptional profile of the Pseudomonas aeruginosa PA14 two-component sensor kinase PA4398 was investigated under swarming conditions using DNA microarrays. To this aim three independent cultures of the PA14 wild-type and the PA4398 mutant were grown until mid-log phase in Luria-Bertani broth following an incubation on BM2-swarm plates containing 0.1 % (wt/vol) casaminoacids and 0.5 % (wt/vol) agar for 20 h at 37 °C. Subsequent total RNA was extracted from the leading edge of dendritic swarm colonies and analyzed by microarrays.