Home Builder Developer - Interior Renovation and Design

When it comes to transforming your backyard, there are few projects bigger than adding a garden fence. In addition to being a simple way to keep your plants guarded from roving wildlife, they can also create a comfortably secluded place in your backyard that you can enjoy in peace. But before you start putting up posts, there are a few tips you'll want to consider to ensure you get the outcome you want. Read on for garden fence ideas that will protect your space and still look great.

RELATED: 9 Essential Gardening Tools Everyone Should Have at Home.

Putting up any kind of fence in your yard is going to catch the eye, which makes the material you use an important decision. According to Marek Bowers, founder of Bolder Green, using reclaimed wood can result in a unique-looking fence that is also sustainable.

"Each plank will have its own character, adding a rustic charm to your yard or garden," he says. "Just ensure the wood is treated and sanded for safety and durability. The weathered look can be incredibly appealing, but you can also paint it to match your garden's aesthetic."

He says you can find reclaimed wood at salvage yards and demolition sites (with permission, of course) or websites such as Etsy, Craigslist, and Facebook Marketplace. You can also reach out to specialty reclaimed wood suppliers and local farms or ranches for anything they might be offloading.

Adding a wooden or stone wall may interrupt the flow of your green space. That's where floral vines can come in handy.

"A bespoke ipe fence adorned with Bignonia capreolatacommonly known as crossvinenot only enhances the aesthetic appeal but also serves as a habitat for hummingbirds, bees, and butterflies," says Amy Hovis, owner of Eden Garden Design. "This crossvine is a fast-growing, evergreen vine that produces striking tubular orange flowers."

She adds that once you're ready to start planting vines, you should install a wire, string, or steel cable to allow the vines to shoot upwards as they begin to grow.

RELATED: Gardening Influencer Reveals the #1 Plant to Give Your Yard Beautiful Color.

Your garden is a place where greenery is meant to thriveand not just in the ground. If you want to create a feeling of lushness that embraces nature, even with your barriers, consider a living wall.

"These fences are made of plant panels that provide privacy and security while adding a lush, green element to your yard, transforming a plain fence into a vibrant backdrop," says Bowers. "Living walls are typically mounted on an existing fence or a sturdy support structure to ensure stability and proper irrigation."

This dense foliage not only enhances privacy by blocking views from neighbors and passersby but also helps absorb sound, making your yard a quieter, more peaceful place.

Not all fences are made of solid wood. If you're looking for an easy way to customize the look and give it some life without installing panels, you can use other materials that help plants grow. ae0fcc31ae342fd3a1346ebb1f342fcb

"We love to use fencing with perforated steel or wire screens to grow vines on," Hovis tells Best Life. "Hedging designs can often be achieved by layering plants of varying heights and textures. This creates visual depth and sensory intrigue while offering an organic look that provides privacy to one's home and landscape."

However, she clarifies that it's important to consider whether the plants you're using are evergreen or deciduous.

"If they're deciduous, plant variety and placement have increased importance so that seasonal coverage remains consistent overall," she explains.

RELATED: 6 Ways to Make Your Lawn Maintenance-Free.

Sturdy wood is an appealing option for anyone truly concerned about security. But there's another simple and affordable material that can look even better for a fraction of the price.

"Bamboo fencing is a fantastic option for those who want an exotic, eco-friendly solution," says Bowers. "It's incredibly strong and grows quickly, making it a sustainable choice. And natural bamboo's light color adds a warm, tropical feel to your garden."

However, it still requires a little work beyond planting or placing: Bowers warns it's essential to secure the bamboo poles tightly and treat them to withstand weather conditions.

Before you start putting posts into the ground, consider yet another living option: For a natural, green barrier, Bowers suggests removing your fence and planting trees tightly together to grow a privacy hedge.

"These are my personal favorites. I actually tear out fences to build these!" Bowers tells Best Life.

Opting for living barriers can can add several important elements to your garden. "This lush, living wall provides excellent privacy and elegance," he says. "Regular trimming will maintain the desired shape and size. But the foliage looks beautiful and absorbs noise, making your yard more serene and pleasant."

See the rest here:
6 Garden Fence Ideas That Protect Your Space and Look Great - Best Life

Natural Resources and Environment portfolio spokesperson Cr Jason Bartels at the soon-to-be-upgraded Laurisen Park in Kepnock.

Safe and diverse play opportunities, from fully-fenced playgrounds, shaded parks and updated skate facilities, are among the essential services prioritised in Bundaberg Regional Councils 2024-25 budget.

In the upcoming financial year, Bundaberg Regional Council will continue to place an emphasis on the enhancement of local recreational areas.

One of the key projects is the refurbishment of Laurisen Park, a highly popular space identified for upgrade in the Parks and Open Space Strategy 2019 2026.

Natural Resources and Environment portfolio spokesperson Cr Jason Bartels said the park, which served the Kalkie and Kepnock areas, would see significant improvements to meet the community's needs and ensure the safety and enjoyment of its users.

A strategic relocation of the play equipment in Laurisen Park will better maximise the site's potential, Cr Bartels said.

While this will move the play area closer to the road the design also includes a barrier fence which will see the playground fully enclosed.

Its the latest in a series of parks to be upgraded with barrier fencing because the community has told us this is a priority to increase accessibility and we are listening.

Additional funding has been incorporated into the budget to allow Council to consider other areas for full fencing in upcoming upgrades.

During a community consultation for the Lake Ellen Heritage Hub and Playground the need for fully enclosed play spaces was identified, he said.

Detailed designs for this project are currently underway, promising a safer and more inclusive environment for all users.

Funding has also been committed to a Skatepark Refurbishment Program which identified Innes Park Reserve Skate Park and Jack Strathdee Memorial Park Skate Park for upgrades based on a comprehensive audit.

Cr Bartels said the program would ensure these popular facilities remained safe and enjoyable for users.

He said visitors to regional parks would also be kept safe from the sun, with Councils Shade Sail Program making strides in improving comfort and usability in several locations.

Shade structures are currently being fabricated for Apple Tree Creek, with Gorman Park, Burnett Shores Park, and Innes Park Reserve scheduled for installations before summer, he said.

Additionally, three shelters are ready for installation at Elliott Heads Skate Park.

Cr Bartels said overall the upgrades reflected Council's dedication to meeting the needs of the community and providing high-quality recreational facilities for all.

View the full Bundaberg Regional Council 2024-25 budget document online.

See original here:
Fences and shading for region's parks Bundaberg Now - Bundaberg Now

Transgenic mice

All experimental procedures for this study were performed at the Biomedical Center, LMU Munich, in accordance with German and European Union guidelines and were approved by the government of Upper Bavaria. Primary cultures of mouse astrocyte were obtained from the cortex of R26-M2rtTA and Yy1tm2Yshi (ref. 61) mice of P56 days of age. R26-M2rtTA (no. 006965) and Yy1tm2Yshi (no. 014649) mice were obtained from The Jackson Laboratory. The mice were not selected based on their gender. The mice were fed ab libitum; housed in individually ventilated cage systems in a room with a temperature of 22C2C, 55%10% humidity and a 12-h/12-h light/dark cycle; and maintained under specific pathogen-free conditions.

Astrocytes were isolated4,32 by dissecting three postnatal mice (P56), and both the gray and white matter of the cerebral cortex were isolated, after removing the subventricular zone, striatum and hippocampus. The cortical meninges were also removed. The cortical tissue was mechanically dissociated, and the cell suspension was centrifuged at 300g, 4C, for 5min. The cell pellet was resuspended in astrocyte medium consisting of DMEM/F12 (1:1) with GlutaMAX (Thermo Fisher Scientific), 10% FBS, penicillinstreptomycin (Gibco), glucose (Gibco), 1 B27 serum-free supplement (Gibco), 10ngml1 epidermal growth factor (EGF, Gibco) and 10ngml1 basic fibroblast growth factor (bFGF, Gibco). The resulting cell suspension was plated onto a T-25 flask. The primary astrocyte culture was maintained in an incubator for 7d at 37C and 5% CO2. Thereafter, the cells were passaged using 0.05% trypsin/EDTA (Thermo Fisher Scientific) and plated onto the following poly-d-lysine (PDL) (Sigma-Aldrich) coated surfaces for the following experiments: 50,000 cells per well in a 24-well plate in 500l of media for immunocytochemistry; 200,000 cells per six-well plate for bulk-RNA-seq, bulk-ATAC-seq, 10x multiome and 10x single-cell RNA-seq experiments; and 1,000,000 cells per T-25 flask for ChIP-seq.

The plasmid FUW-TetON was modified to insert Gateway cloning sites. Mouse Ngn2, eGFP and 9S-A Ngn2 (referred to as PmutNgn2, which was a gift from A. Philpott)25 were cloned into the Gateway entry vectors (Thermo Fisher Scientific) and subsequently shuttled into the dox-inducible lentiviral expression vector FUW-TetON by employing Gateway recombination cloning technology (Thermo Fisher Scientific). The lentiviral expression vector was characterized by the presence of a tetracycline response element followed by the mammalian CMV2 promoter, which regulated the expression of the TFs and the eGFP (fluorescent reporter employed to identify transduced cells). The TF sequence was separated from the eGFP sequence by an internal ribosome entry site (IRES).

Vesicular stomatitis virus-glycoprotein (VSV-G)-pseudotyped lentiviral particles were produced by transfecting 293T cell line with the following plasmids: pCMVdR8.91 (expressing gag, pol and rev genes), pVSVG and lentiviral expression plasmid. The lentiviral particles were harvested and concentrated by ultracentrifugation at 125,000g for 2h, and the pellet containing the lentiviral particles was resuspended in 1 PBS (supplemented with 5mM MgCl2). The lentivirus was aliquoted and stored at 80C until use. The lentiviral titer was determined by a functional assay, where primary mouse astrocytes were infected with the lentivirus preparation at various dilutions, and the number of successfully infected cells was determined by immunostaining the transduced cells with an anti-GFP antibody (for TF-encoding lentiviruses) or an anti-RFP antibody (for Cre-expressing lentivirus). The viral titers used in all the experiments were in the range of 1010 to 1012 transducing units per milliliter.

After seeding the desired number of cells in PDL-coated plates, 24h later the cells were transduced with 107 to 109 transducing units per microliter of lentiviral particles. Approximately 20h after transduction, the astrocyte medium was replaced with fresh medium containing DMEM/F12 (1:1), supplemented with penicillinstreptomycin, glucose, 1 B27 and GlutaMAX (differentiation medium), and the cells were maintained in culture in a 9% CO2 incubator for a period, depending upon the experimental design. To induce the expression of the TF and fluorescent protein, dox (2gml1) was added to the differentiation medium, and the dox-containing medium was added freshly for four consecutive days.

Cells were prepared for fluorescence-activated cell sorting (FACS) by washing them once with 1 PBS followed by trypsinization (0.05% trypsin in EDTA) for 5min. The trypsinization reaction was stopped by adding astrocyte medium. The harvested cells were then washed twice with ice-cold PBS and centrifuged at 300g for 3min at 4C. The cells were resuspended in DMEM/F-12 (1:1), and a single-cell suspension was generated using a 40-m cell strainer. FACS was performed by employing a FACSAria Fusion (BD Biosciences) using a 100-m nozzle. The gating strategy was set by using forward, side scatter and untransduced astrocytes as a negative control and eGFP-expressing astrocytes as a positive control. Additionally, for Methly-HiC, astrocytes were stained for DAPI, and only cells in G0 and G1 (single DNA content) were sorted. The cells were sorted into DMEM/F-12 (1:1).

Coverslips containing astrocytes were fixed using 4% paraformaldehyde in 1 PBS for 10min at room temperature. The cells were washed twice with 1 PBS and stored for up to 3weeks at 4C before staining. The coverslips were incubated with blocking solution (3% BSA, 0.5% Triton X-100 in 1 PBS) for 30min. Thereafter, the coverslips were incubated with the primary antibody diluted (for detailed information about antibodies used, see Supplementary Table 4) using blocking solution overnight at 4C. After washing the coverslips three times with 1 PBS, they were incubated with the appropriate secondary antibody (diluted 1:500) for 1h at room temperature. The coverslips were stained with DAPI (diluted 1:1,000 in blocking solution) for 10min at room temperature. Finally, the coverslips were mounted using Aqua-Poly/Mount (Polysciences).

A Zeiss Cell Observer was employed to perform continuous live imaging of astrocyte-to-neuron conversion. The acquisition of images was performed as follows. Phase contrast images and fluorescent images (GFP) were captured every 20min and 4h, respectively, with a 10 phase contrast objective (Zeiss) and an AxioCam HRm camera. Zeiss AxioVision 4.7 software was controlled by a custom-made VBA module (TAT, Timm Schroeder, ETH Zrich)62. The movie processing and analysis was performed in ImageJ (1.53q) (National Institutes of Health).

The acquisition of microscopy images was performed using an AxioM2 epifluorescence microscope (Zeiss) or an LSM 710 laser scanning confocal microscope (Zeiss) and ZEN2 software (version 2.0.0.0, Zeiss). The quantification of iNs was performed by applying the following stringent criteria, which were previously described in Gascon et al.5. iNs had to possess a unipolar or bipolar morphology, with a process being at least three times the length of its soma. Additionally, the iNs had to be III-tubulin positive and GFAP negative. In case of the live-imaging microscopy, the time of conversion was defined as a timepoint (in hours) when a GFP+ cell acquired neuronal morphologythat is, exhibited a unipolar or bipolar morphology where the process was at least three times the length of its soma. Statistical analysis was performed in R (version 4.2.1). In Figs. 1eh,j and 7c and Extended Data Fig. 1e, statistical significance was calculated with linear regression by implementing the function lm in RStudio on log2-transformed reprogramming rate14.

The primary astrocytes, transduced with the GFP, Ngn2 or PmutNgn2 lentivirus, were obtained from the same litter of mice. In case of the primary astrocytes obtained from the Yy1tm2Yshi line for the functional studies (conditional knockouts of the candidate gene, Yy1), the wild-type, heterozygote and homozygote genotypes were obtained from same litter of mice by crossing two heterozygote mice.

No statistical methods were used to pre-determine samples sizes, but our sample sizes relied on previous experience, showing that this sample size gives sufficient statistical power5,6,17,18,19,45,63. No data were excluded from the analyses. For data in Figs. 1eh,j and 7c, the values were log transformed and, hence, assumed to be normally distributed.

All the data analysis for immunocytochemistry (Figs. 1eh and 7c) and live imaging (Fig. 1j) was blinded. The genomic experiments and associated data analysis were not blinded because they did not involve subjective measurements.

For the Methyl-HiC experiment, the cells were stained with DAPI following the intracellular staining protocol with the following modifications19. Upon fixing with 1% formaldehyde and permeabilizing the cells, they were stained with DAPI (1:1,000 dilution in wash buffer containing 1% BSA, 0.1% RNasin plus RNase inhibitor (Promega) in PBS). The cells were washed once with the wash buffer and subsequently resuspended in PBS with 1% BSA and 1% RNasin plus RNase inhibitor, filtered through a 40-m cell strainer and FACS sorted.

Approximately 30,000 events per condition were FACS sorted into DMEM/F-12 (1:1) and centrifuged at 300g for 5min at 4C. Then, the cell pellet was resuspended in TRIzol (Thermo Fisher Scientific) and further processed with an RNA Clean & Concentrator Kit (Zymo Research) to extract the RNA. The quality of the extracted RNA was determined using an Agilent RNA 6000 Pico Kit and an Agilent 2100 Bioanalyzer system. All the samples used for library preparation had an RNA integrity number (RIN) value>8.

Next, 50ng of RNA was used as the input material for library generation, and the protocol was a bulk adapted version of mcSCRB-seq64,65. cDNA was generated from the poly(A)-enriched RNA fraction using oligo-dT primers and a Maxima First Strand cDNA Synthesis Kit (Thermo Fisher Scientific). The unincorporated primers were digested using Exonuclease I (Thermo Fisher Scientific). The resulting cDNA was pre-amplified using Terra polymerase (Takara Bio). The quality of the cDNA was determined using the Agilent 2100 Bioanalyzer system. The RNA-seq library was prepared using a NEBNext Ultra II FS DNA Library Kit for Illumina (New England Biolabs) according to the manufacturers instructions. The quality of the RNA-seq libraries was assessed using the Agilent 2100 Bioanalyzer system.

Bulk ATAC-seq libraries were generated by following the OMNI-ATAC-seq protocol66. Approximately 70,000 events were FACS sorted into tubes containing DMEM/F-12 (1:1) and centrifuged at 300g for 5min at 4C, and the cell pellet was resuspended in ATAC resuspension buffer. The cell viability and cell number were determined using a Countess automated cell counter (Thermo Fisher Scientific). Fifty thousand viable cells were used for the Tn5 transposition reaction. The transposition reaction was performed at 37C for 30min in an Eppendorf thermomixer. The transposed fragments were purified using a DNA Clean & Concentrator-5 Kit (Zymo Research). The purified transposed DNA fragments were amplified using NEBNext Ultra II Q5 Master Mix (New England Biolabs) and cleaned up using the DNA Clean & Concentrator-5 Kit. The quality of the ATAC-seq libraries was assessed using the Agilent 2100 Bioanalyzer system.

The ChIP-seq protocol was adapted from a previously described protocol67. In brief, 4 million astrocytes were fixed using 1% methanol-free formaldehyde (Thermo Fisher Scientific) at room temperature for 10min. The cross-linking reaction was terminated by the addition of 125mM glycine followed by an incubation step at room temperature for 5min. The cells were lysed by suspension in a hypotonic buffer (20mM Tris, pH 7.4; 2mM MgCl2; 5% glycerol; 0.6% NP-40) and incubation on ice for 5min with mild vortexing every 30s, which resulted in the release of the nuclei. The nuclei were resuspended in ChIP lysis buffer (20mM Tris, pH 7.4; 150mM NaCl; 1% sodium deoxycholate; 0.1% SDS; 1mM EDTA, pH 8.0) and sonicated using a Bioruptor Pico sonicator (Diagenode) with the following settings: 30s ON/OFF, 20 cycles. The sonicated chromatin was quality controlled using the Agilent 2100 Bioanalyzer system. The sonicated chromatin used for ChIP-seq ranged from 150bp to 300bp.

The chromatin was pre-cleared using Dynabeads Protein G (Thermo Fisher Scientific). After pre-clearing, 10% of the pre-cleared chromatin was set aside as the input fraction. The chromatin was incubated with 4g of mouse monoclonal anti-FLAG M2 antibody (Sigma-Aldrich) overnight at 4C on a rotating wheel (10r.p.m.). After the ChIP, Dynabeads Protein G (Thermo Fisher Scientific) was added to the ChIP sample and incubated at 4C for 3h on a rotating wheel (10r.p.m.). The ChIP sample was washed five times with LiCl was buffer (50mM Tris, pH 7.4; 1mM EDTA, pH 8; 1% NP-40; 1% sodium deoxycholate; 0.5M LiCl) followed by a single wash with TE buffer (10mM Tris, pH 8; 1mM EDTA, pH 8). All the wash steps were performed for 5min at 4C on a rotating wheel (10r.p.m.). The elution of the proteinDNA complex was performed using the elution buffer (50mM NaHCO3, 1% SDS) under the following condition: constant agitation on a thermomixer (Eppendorf) at 60g for 15min at 65C. The eluted DNA was de-crosslinked by the addition of 5M NaCl (final concentration: 210mM) and incubated overnight (not more than 15h) at 65C.

The de-crosslinked DNA was treated with RNase A (Thermo Fisher Scientific) and incubated in a thermomixer (Eppendorf) at 60g for 90min at 37C, followed by treatment with Proteinase K (Ambion) and incubated in a thermomixer (Eppendorf) at 800r.p.m. for 120min at 55C. The DNA was extracted using UltraPure Phenol:Choloroform:Isoamylalcohol (25:24:1, v/v, Thermo Fisher Scientific) following the manufacturers instructions and precipitated by ethanol precipitation (glycogen, 3M sodium acetate, pH 5.2, 100% ethanol) overnight at 20C. The DNA was resuspended in low TE buffer and quantified Qubit dsDNA HS (Thermo Fisher Scientific). One nanogram of ChIP DNA was used as starting material for library preparation with the MicroPlex Library Preparation Kit v2 (Diagenode). The quality of the ATAC-seq libraries was assessed using the Agilent 2100 Bioanalyzer system.

The Yy1, FLAG, Rad21 and H3K27Ac CUT&RUN assays were performed as previously described with specific modifications63.

In brief, 23.6105 iNs were harvested, washed twice and resuspended in wash buffer (20mM HEPES, pH 7.5; 150mM NaCl; 0.5mM spermidine; 1 Roche cOmplete). Concavalin A beads (BioMag Plus, Polysciences) were activated with bead activation buffer (20mM HEPES, pH 7.9; 10mM KCl; 1mM CaCl2; 1mM MnCl2). Cells were incubated with 10l of activated beads for 10min at room temperature. After incubation, the beads were resuspended in a cold antibody buffer (2mM EDTA in digitonin buffer) containing antibody (5g of Yy1 (D5D9Z) rabbit monoclonal antibody 46395, 2g of FLAG antibody (Sigma-Aldrich, F3165-.2MG), 5g of Rad21 (BIOZOL, GTX106012) and 1g of H3k27Ac (Abcam, 39133)), and the mixture was incubated on a nutator overnight at 4C.

On the next day, the beads were washed twice and resuspended in 0.75l of pAG-Mnase in digitonin buffer (0.1% digitonin, Thermo Fisher Scientific, in wash buffer) and incubated for 10min at room temperature on a rotator. Later, beads were washed twice with cold digitonin buffer and then resuspended in 50l of digitonin buffer containing 1l of 100mM CaCl2. The suspension was incubated for 2h at 4C on a nutator. After the incubation, 33l of STOP buffer (340mM NaCl, 20mM EDTA, 4mM EGTA, 50gml1 RNase A (Thermo Fisher Scientific), 50gml1 glycogen) was added to each reaction, and the mixture was incubated for 30min at 37C.

DNA extraction was performed using UltraPure Phenol:Chloroform:Isoamyl Alcohol (25:24:1, v/v, Thermo Fisher Scientific) and precipitated with 100% ethanol, 1l of glycogen and 1/10th volume of 3M sodium acetate for 416h at 20C. DNA was then dissolved in 10l of 1mM Tris-HCl, pH 8, and 0.1mM EDTA.

CUT&RUN libraries were prepared with an NEBNext Ultra II DNA Library Prep Kit for Illumina using 630ng of fragmented DNA. The quality of the CUT&RUN libraries was evaluated using the Agilent 2100 Bioanalyzer system.

Single-cell multiome (version 1, 10x Genomics) libraries were generated according to the manufacturers instruction manual. In case of the multiome libraries, we targeted for the recovery of 500 nuclei for the GFP, Ngn2 and PmutNgn2 conditions and 5,000 nuclei for the Astro condition.

A modified Methyl-HiC was performed19 based on previously described protocols17,18. Full details of the experimental steps can be found at https://www.protocols.io/view/methylhic-bif2kbqe/.

Pellets from frozen, fixed and FACS-sorted G0/G1 cells were thawed and then lysed on ice with 0.2% Igepal-CA630 (Sigma-Aldrich). Nuclei were subsequently permeabilized with 0.5% SDS and chromatin digested with DpnII (New England Biolabs) at 37C overnight. DpnII was heat inactivated at 62C, and then sticky ends were filled in with biotin-14-dATP (Life Technologies) before proximity ligation with T4 Ligase (New England Biolabs). Proteinase K (New England Biolabs) and NaCl was used for reverse crosslinking nuclei overnight at 68C, and DNA was afterward purified using ethanol precipitation. A Covaris S220 sonicator was next used to shear the DNA to approximately 550-bp fragments.

End repair was performed on the sonicated DNA with T4 DNA Polymerase (New England Biolabs). Approximately 0.01% of methylation controls were spiked into sample, and the reaction was bisulphite converted using an EZ DNA Methylation-Gold Kit (Zymo Research). Libraries were prepared using an Accel-NGS Methyl-Seq DNA Library kit (Swift Biosciences) according to the manufacturers instructions until the adaptor ligation step. At this point, streptavidin T1 beads (Thermo Fisher Scientific) were used for biotin pulldown of DNA, followed by stringent washes. Final libraries were amplified from the streptavidin beads using EpiMark Hot Start Taq (New England Biolabs) with Methyl-Seq indexing primers (Swift Biosciences), followed by size selection with 0.6 AMPure XP beads (Agencourt).

P19 cells were plated in 10-cm dishes. Cells were transfected using Lipofectamine 3000 with 5g of Control (Pcig2), Neurogenin2 and Neurogenin2 mutated (S-A9 TA1) DNAs and were harvested after 24h by cell scraping using cold PBS followed by centrifugation at 300g for 5min to collect the cell pellets.

The P19 cell pellets were thawed on ice and resuspended in 1 pelleted cell volume of the lysis buffer A (10mM HEPES, pH 7.9; 1.5mM MgCl2; 10mM KCl; 0.1% NP40; 1 protease inhibitor cocktail (Roche, 04 693 116 001); 50mM sodium fluoride; 0.2mM sodium orthovanadate; 0.05mM MG132 (Sigma-Aldrich, M7449); 1mM PMSF). After leaving the resuspended cells for 5min on ice, an equal volume of lysis buffer B (10mM HEPES, pH 7.9; 1.5mM MgCl2; 10mM KCl; 0.1% NP40; 1 protease inhibitor cocktail (Roche); 50mM sodium fluoride; 0.2mM sodium orthovanadate; 0.05mM MG132 (Sigma-Aldrich, M7449); 1mM PMSF) was added to leave another 5min on ice. Cells were lysed by pipetting up and down followed by passing through a 27.5-gauge needle (insulin syringe) for 1012 times on ice. This was followed by centrifugation at 15,000g for 15min, and the supernatant was collected. For in vivo samples, embryonic cortex (dorsal telencephalon) was collected at E12.5 and E14.5 to proceed with protein extraction as above.

IP was performed using 2g of anti-YY1 antibody (mouse anti-YY1; Santa Cruz Biotechnology, sc-7341) and control mouse IgG from in vivo (embryonic cortex) and in vitro (P19 cells) samples. Anti-YY1 antibody was incubated with Protein G Magnetic Dynabeads at 4C for 13h in IP 150 KCl buffer (25mM Tris, pH 7.9; 5mM MgCl2; 10% glycerol; 150mM KCl; 0.1% NP40; 0.3mM DTT; 1 protease inhibitor cocktail (Roche, 04 693 116 001), 50mM sodium fluoride; 0.2mM sodium orthovanadate; 0.05mM MG132 (Sigma-Aldrich, M7449); 1mM PMSF). Then, 0.05% NP40 was added to the protein and centrifuged at 17,530g for 15min. The supernatant was collected and added with 0.1mgml1 ethidium bromide to incubate for 30min, followed by centrifugation at 17,530g for 15min. The supernatant was then pre-cleared with Protein G Dynabeads for 1h by end-over-end rotation at 4C. After pre-clearing, protein was added to the Dynabeads, which were previously incubated with anti-YY1 antibody, followed by overnight rotation at 4C. The supernatant was removed after overnight incubation, followed by four washes using PBS with protease inhibitors (0.3mM DTT; 1 protease inhibitor cocktail (Roche, 04 693 116 001); 50mM sodium fluoride; 0.2mM sodium orthovanadate; 0.05mM MG132 (Sigma-Aldrich, M7449); 1mM PMSF). The proteins bound to the beads were eluted using 2 Laemmli buffer, by heating at 95C for 5min. Proteins were separated from beads using a magnet and proceeded to western blotting to visualize the immunoprecipitated proteins.

The immunoprecipitated proteins were run on 12% SDS-PAGE gels at 70V during stacking and 120V while resolving. The proteins were transferred to PVDF membranes (1620177, Bio-Rad) in transfer buffer (25mM Tris; 192mM glycine; 20% methanol, pH 8.3) at 40V overnight at 4C after the SDS-PAGE. Membranes were blocked in TBST (10mM Tris; 100mM NaCl, pH 7.4; 0.1% Tween 20) with 5% (w/v) skim milk for 1h at room temperature and then incubated with primary antibodies overnight at 4C. Membranes were washed 310min in TBST and then incubated for 1h at room temperature with 1/50,000 dilutions of horseradish peroxidase (HRP)-coupled secondary antibodies (anti-rabbit IgG, 7074S, Cell Signaling Technology). Membranes were washed 310min at room temperature and then processed with ECL Plus Western Blotting Reagent (29018904, GE Healthcare) before developing with X-ray film (1141J52, LabForce) and a Bio-Rad ChemiDoc MP Imaging System. The primary antibodies used were rabbit anti-YY1 (Invitrogen, MA5-32052), rabbit anti-Neurogenin2 (Invitrogen, PA5-78556) and rabbit anti-Ezh2 (Cell Signaling Technology, 5246).

Single-cell multiome reads were aligned to the Mus musculus reference genome (GRCm38, mm10), and the quantification was performed using cellranger-arc-2.0.1. Data were analyzed using Signac (version 1.7.0)68 and ArchR44. The quality control (QC) metrics are reported in Supplementary Table 1.

We eliminated low-information content cells based on the following selection criteria: cells where fewer than 1,000 genes and 1,000 unique molecular identifiers (UMIs) (from the gene expression library) and fewer than 8,000 unique fragments per cell, transcription start site (TSS) enrichment <1 and nucleosome signal <0.2 (from the ATAC library) were detected. To avoid including possible doublets in the further analysis, cells where more than 30,000 genes (from the gene expression library) and more than 125,000 unique fragments, TSS enrichment >20 and nucleosome signal >2 (from the ATAC library) were eliminated. Nucleosome signal and TSS enrichment were calculated using Signac (version 1.7.0)68 and plotted using ggplot2. Fragment lengths were calculated using ArchR44 and plotter using ggplot2. Upon filtering out the low-quality cells from all the conditions, the number of cells from the Astro condition was balanced with the other conditions.

The individual modalities (gene expression and ATAC) were normalized and processed using Signac68 and Seurat (version 4.0)33. In brief, peak calling was performed on pseudobulk aggregate per condition using MACS2. A high-quality union peak set was identified by merging the individual peaks and filtering out peaks, which overlapped with a list of blacklisted regions. The count matrix for the high-quality peak set was generated and incorporated into a Seurat object. It was subjected to TF-IDF normalization followed by SVD as described previously. For the gene expression modality, after log transformation, variance-stabilizing transformation was used to perform feature selection. Principal component analysis was performed using the first 20 dimensions. We then computed a joint neighbor graph that represents both gene expression and chromatin accessibility using FindMultiModalNeighbors. We then applied Louvain clustering to cluster cells (resolution=0.2, n.start=20, n.iter=30, algorithm=1), and the cell clusters were visualized using UMAP (min.dist=0.5, spread=1.5, n.components=2L). Cluster identity was determined based on the top 40 differentially expressed genes (MAST, minimum expression change of 0.25 and expressed by at least 25% of the cells in the cluster)69 as well as known marker genes.

Maturation pseudotime analysis was implemented on the QC-approved cells using Monocle3 (refs. 34,35,36,37). The UMAP coordinates was retained from Seurat and used to build the cds object in Monocle3. Cells in cluster iN_1 were selected as the root cells, and a trajectory graph was constructed using the following parameters: minimal_branch_len=5, maxiter=30. The change in gene expression along the constructed trajectory was calculated by fitting a generalized additive model employing cubic regression splines and REML smoothing. The resulting values were rescaled from 0 to 1.

The calculation of motif accessibility deviation scores using position weight matrices obtained from the JASPAR2000 database and Ngn2 ChIP-seq was performed as described previously19 using the ChromVar implementation in Signac68. TF footprints were calculated using ArchR44 and visualized using ggplot2.

To link putative enhancers with their target genes, we used ArchR with empirical P value estimation and k=50. We distinguish among positively correlated (r>0.35; false discovery rate (FDR)<0.1), negatively correlated (r<0.35; FDR<0.1) and non-correlated pairs (0.35

We reasoned that we can predict direct targets of a TF either by using available ChIP-seq peaks or based on the enrichment of the TF motif in the positively correlated EGPs. First, we identified all EGPs that contained the corresponding ChIP-seq peak or TF motif (either in the distal region or in the promoter region). Thereafter, we calculated the gene linkage score by adding up the r2 from each pair per gene (if the peak/motif was contained in the promoter, we used a value of r=1). To calculate enrichment, we used background ATAC peaks with similar GC content and determined significance using a hypergeometric test. A potential limitation of this method is that the significance of peak/motif enrichment for genes with very few identified pairs cannot be accurately calculated.

The experimental conditions were labeled according to the manufacturers instructions with the following CellPlex reagents from the 3 CellPlex Kit set A (10x Genomics, PN:1000261): Yy1 WT (CMO309), Yy1 KO (CMO310), Yy1 WT/Ngn2+ (CMO311) and Yy1 KO/Ngn2+ (CMO312). Approximately 25,000 events per condition were FACS sorted (Yy1 WT; untransduced, Yy1 KO; RFP+, Yy1 WT/Ngn2+; GFP+, Yy1 KO/Ngn2+; RFP+GFP+) into an Eppendorf tube. Approximately 33,000 cells were loaded onto a Chromium Next GEM ChIP G (10x Genomics, PN:2000177) to obtain a targeted cell recovery of 20,000 cells. The gene expression library (PN:3000431, single cell 3 v3) and the cell multiplexing library (PN:3000482) were prepared according to the manufacturers protocol (CG000388, Rev A). The gene expression library and the cell multiplexing library were quality controlled using the Agilent 2100 Bioanalyzer, and the libraries were sequenced according to the manufacturers specifications.

Single-cell RNA-seq reads were aligned to the Mus musculus reference genome (GRCm38, mm10), and the sample assignment and quantification were performed using cell ranger multi in cellranger-6.0.0. The QC metrics are reported in Supplementary Table 1.

We eliminated low-information content cells based on the following selection criteria: cells where fewer than 1,000 genes and 2,500 UMIs were detected. To exclude dead cells, we filtered out cells containing more than 20% mitochondrial reads. To avoid including doublets in the further analysis, cells containing more than 6,000 genes were excluded.

Seurat (version 4.0) was used to analyze the cells that passed the filtering steps. The data were normalized using SCTransform, and principal component analysis was performed using the first 25 dimensions. We applied Louvain clustering (resolution=0.6, n.start=20, n.iter=20), and the data were visualized by UMAP projection (min.dist=0.5, spread=1.5, n.components=2L). Cluster identity was determined based on the top 40 differentially expressed genes (MAST, minimum expression change of 0.25 and expressed by at least 25% of the cells in the cluster)69.

The ATAC-seq FASTQ files were demultiplexed using Je (version 1.2)70, and the demultiplexed reads were aligned to the mouse genome (GRCm38, mm10). Post-alignment read filtering, peak calling and irreproducible discovery rate (IDR)-based peak filtering were performed by implementing the ENCODE ATAC-seq pipeline. The sequencing and QC metrics are listed in the form of a supplementary data table. The bigWig coverage track was generated using deepTools (version 3.1.3)71. The plotting of the ATAC-seq signal at genomic features was performed using SeqPlots72. The QC metrics are reported in Supplementary Table 1.

The RNA-seq FASTQ files were demultiplexed using Je (version 1.2)70; demultiplexed reads were aligned to the mouse genome (GRCm 38, mm10) using STAR (version 2.7.1a)73; and read counts per gene were obtained by using the quantMode GeneCounts option. Further analysis was performed using DEseq2 (ref. 74) in RStudio. The result table for pairwise comparison between PmutNgn2 versus Ngn2 was used the input to generate the GO term enrichment bubble plot in the R package clusterProfiler75. The QC metrics are reported in Supplementary Table 1.

The ChIP-seq FASTQ files were demultiplexed using Je (version 1.2)70; demultiplexed reads were aligned to the mouse genome (GRCm38, mm10); and post-alignment read filtering, peak calling and IDR-based peak filtering were performed by implementing the ENCODE ChIP-seq pipeline. The bigWig coverage track was generated using deepTools (version 3.1.3)71. The plotting of the ChIP-seq signal at genomic features was performed using the R package SeqPlots72. The QC metrics are reported in Supplementary Table 1.

CUT&RUN data were uniformly processed using CUT&RUN tools 2.0 (ref. 76). Peaks were called using MACS2, and the bigWig coverage track was generated using deepTools (version 3.1.3)71. The QC metrics are reported in Supplementary Table 1.

FASTQ files from the Methyl-HiC were mapped to the mouse genome (GRCm38, mm10) by employing JuiceMe77. Further analysis was performed only with uniquely mapping reads (mapq score>30). After the elimination of polymerase chain reaction (PCR) duplicates, the translation of reads into a pair of fragment ends (fends) was achieved by the association of each read with its downstream fend. MethylDackel was used to assess CpG methylation, which entailed the elimination of the initial six nucleotides in the mergeContext mode. Pooling of reads from individual replicates was performed, and, for a cytosine to be considered for further analysis, it had to be in the CpG context and possess at least 10 total coverage. In case of Hi-C, exclusion of reads was based on the following criteria: mapped to the same restriction fragment and separated by less than 1kb. The QC metrics are reported in Supplementary Table 1.

Filtered fend-transformed read pairs were imported into the following genome database: mm10 after conversion into misha tracks. The Shaman package was used for read pair normalization (https://tanaylab.bitbucket.io/shaman/index.html), and the calculation of the Hi-C score was performed by employing k-nearest neighbors (kNN)22.

The calculation of contact probabilities as a function of genomic distance was previously described22. The insulation score, which is used to define insulation on the basis of observed contacts, was also previously described 19,22,78, and differential TAD boundaries were identified using insulation score19,22.

The calculation of contact matrices dominant eigenvector, which have been binned at 250kb, was previously described and performed using publicly available scripts (https://github.com/dekkerlab/cworld-dekker)79. Compartment strength was determined by plotting the log2 ratio of observed versus expected contacts (intrachromosomal separated by at least 10Mb) between AA, BB or AB domains. A ratio between the sum of observed contacts within the A and B compartments and the sum of intercompartment contacts was calculated to determine the compartment strength19.

The calculation of insulation and contact enrichment within TADs was previously described19,22.

Two complementary approaches were employed for the calculation of contact enrichment ratio at genomic feature pairs, such as Ngn2 ChIP-seq peaks or EGPs. Aggregated Hi-C maps were used to calculate the log2 ratio of observed versus expected contacts within a specified window size, which was centered on the feature of interest. The average enrichment ratio was also calculated for the following: contact strength in the center of the window versus each of the corners. Furthermore, the extraction of kNN-based Hi-C score for each pair in a 10-kb window centered around it and its representation as a scatter plot or box plots enabled the identification of pair-specific trends. Significance testing was performed by using the Wilcoxon rank-sum test.

Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article.

Here is the original post:
Direct neuronal reprogramming of mouse astrocytes is associated with multiscale epigenome remodeling and requires ... - Nature.com

Image: https://www.getnews.info/wp-content/uploads/2024/07/1720115164.png

Aboveboard Roofing & Remodeling is a leading roofing company. In a recent update, the company shared tips on extending a roof's lifespan. Freeland, MI - In a website post, Aboveboard Roofing & Remodeling shared tips to extend a roof's lifespan.

The roofing contractors Freeland [https://www.aboveboardmi.com/discovering-auburn-mi-a-quaint-gem-in-mid-michigan/] asserted that maintaining a roof is essential to retaining the structural integrity and aesthetic appeal of any home. Frequent inspections are important for identifying and addressing small issues before they escalate. Keeping gutters clear of debris helps prevent water buildup, which can cause leaks and damage. Additionally, trimming overhanging branches minimizes the risk of branches falling and damaging the roof during strong winds.

The experts said that when it comes to roofing installation Freeland [https://www.aboveboardmi.com/discovering-auburn-mi-a-quaint-gem-in-mid-michigan/], proper ventilation is vital in avoiding moisture buildup in the attic, which can cause mold and rot. Installing high-quality underlayment provides an additional layer of protection against water infiltration and improves the roof's durability. Choosing the right roofing material for the climate and environment ensures longevity and reduces the need for frequent repairs.

The experts noted that for roofing replacement Freeland [https://www.google.com/maps?cid=16014189251778533116], timing is crucial. Knowing when it is time to replace a roof can prevent extensive damage to the home's interior and structural integrity. Signs such as curling shingles, missing granules, or visible cracks indicate the need for replacement. Properly preparing the roof and ensuring all old materials are removed before installing new roofing materials is essential for a long-lasting roof.

Location: https://www.google.com/maps/embed?pb=!1m18!1m12!1m3!1d371022.1680275539!2d-84.0325911!3d43.406065950000006!2m3!1f0!2f0!3f0!3m2!1i1024!2i768!4f13.1!3m3!1m2!1s0x8823c9e0ad8cf075%3A0xde3dd447162356fc!2sAboveboard%20Roofing%20%26%20Remodeling!5e0!3m2!1sen!2s!4v1720112944418!5m2!1sen!2s

About Aboveboard Roofing & Remodeling

Aboveboard Roofing & Remodeling [https://www.aboveboardmi.com/] is a premier roofing company. With a focus on professionalism and attention to detail, the firm strives to build lasting relationships with homeowners and businesses alike. It keeps on evolving its services to meet the evolving demands of the market. From initial consultation to project completion, the crew's devotion to quality and client satisfaction shines through in every aspect of its work. Media Contact Company Name: Aboveboard Roofing & Remodeling Contact Person: Taylor Green Email: Send Email [http://www.universalpressrelease.com/?pr=aboveboard-roofing-remodeling-shares-tips-to-extend-a-roofs-lifespan] Phone: (989) 455-3452 Address:7542 Thornberry Dr City: Freeland State: Michigan Country: United States Website: https://www.aboveboardmi.com/

This release was published on openPR.

Go here to see the original:
Aboveboard Roofing & Remodeling Shares Tips to Extend a Roof's Lifespan - openPR

11

Northwest Louisiana Technical Community College (NLTCC) is excited to announce the start of its new Plumbing Program in partnership with the Plumbing-Heating-Cooling Contractors of Louisiana (PHCCLA). The inaugural class of students will begin on August 12th on the NLTCC Shreveport campus and will provide a fantastic opportunity for those interested in a career in plumbing.

The plumbing program is a 48-credit hour technical diploma that combines classroom theory education with hands-on activities. This blend of learning methods ensures students understand the fundamental principles of plumbing while gaining practical skills through lab sessions and on-the-job training with paid apprenticeship opportunities. The curriculum is designed to meet industry standards, ensuring graduates are well-prepared for the workforce. Upon completion of the 48-credit technical diploma and 3,500 apprenticeship on the job training hours, graduates will be eligible to take the Residential Plumber license test. Students can also return to NLTCCs Plumbing year III and IV Workforce Program that will give them the opportunity to prepare for the Journeyman license while completing the remaining 3,500 apprenticeship on the job training hours required for the test.

NLTCC offers various credentials throughout the plumbing program, including:

Construction Laborer, Certificate of Technical Competency, OSHA-10 Certificate Plumber Helper Level I, Certificate of Technical Studies

Plumber Helper Level II, Certificate of Technical Studies

Plumbing, Technical Diploma

A career in plumbing offers numerous benefits, especially in Louisiana, where plumbers are in high demand. In fact, more than 114,500 workers will be needed by 2028. The average technicians salary in Louisiana is more than $54,620 with benefits. Its a stable career with excellent job security, competitive wages, and opportunities for advancement. With the skills and credentials gained from NLTCCs program, graduates will be well-equipped to enter this rewarding field.

If you are interested in learning more about the Plumbing Program at NLTCC or the available financial aid opportunities, visit https://bit.ly/NLTCCPlumbing for more information or call 318-676-7811 to speak to a member of the student services team.

Read more:
NLTCC launches new plumbing program - Minden Press-Herald

A Jewish woman in London was left shocked and upset when a plumber rejected her request for a repair because she did not support the anti-Israel Boycott, Divestment and Sanctions (BDS) movement.

AWG Plumbing and Heating, which is based in Mayfair and should not be confused with AGW Heating, Plumbing and Electrical, based in Hendon, claims online that every member of the company undergoes rigorous training to ensure they meet the highest standards in gas and plumbing services.

But when the woman contacted the firm, a staff member who gave his name as Adam replied by text: I wont be able to provide you any services as were trying to VET all of our customers in the present climate and it appears that you opposed the BDS movement and give cover to the state of Israel as you a part [sic] of the law makers in this country.

"So for this reason, we can no longer offer our services and would therefore no longer like to be contacted."

The BDS campaign, which is modelled after the movement against South African apartheid, was launched in 2005 to end Israels occupation of the Palestinians.

It calls for individuals and businesses to cut ties with the Jewish state and divest from its economy.

After she was denied service because she did not support the campaign, the Jewish customer said: I was extremely shocked and upset. I couldnt believe it had happened to me in this country.

When contacted by the JC, Adam said AWG does not refuse to serve people because they support Israel, but do if they support Zionism or genocide.

While he would not investigate the views of Joe Bloggs, he claimed, he does research customers with a public profile to examine their position on Israel.

We worry that antisemitism and anti-Zionism are being conflated and worry wed be criminalised for our views, Adam said of AWGs staff.

"We have a personal relationship with all our clients, thats one of the great things about interacting with different people.

Lobby group UK Lawyers for Israel (UKLFI) have since reported AWG to the Equalities and Human Rights Commission (EHRC).

They say the company is in breach of section 29 of the Equality Act 2020, which prohibits firms from discriminating against potential customers on the basis of protected characteristics that include religion and philosophical belief.

"AWG discriminated against the customer on the grounds that she supported the State of Israel and opposed BDS. In other words, because she is a Zionist, UKLFI director Caroline Turner wrote.

"In addition, since a higher number of Jews than of other religious groups are Zionists, AWG was also indirectly discriminating against people of the Jewish religion.

Therefore AWG is demonstrating discrimination against Jews and Zionists.

See more here:
Meet the London plumber who will not unblock your toilet if you are a Zionist - The Jewish Chronicle

A Jewish woman in London was left "shocked and upset" when an antisemitic plumber refused her request for work at her home because she supports Israel.

The plumber works at AWG Plumbing and Heating, a company based in London's upscale Mayfair district. The company's website states that all its workers undergo "rigorous training to ensure they meet the highest standards in gas and plumbing services."

However, when the woman contacted the company, she received a text message reading, I wont be able to provide you any services as were trying to VET all of our customers in the present climate and it appears that you opposed the BDS movement and give cover to the state of Israel as you a part [sic] of the law makers in this country. So for this reason, we can no longer offer our services and would therefore no longer like to be contacted."

"I was extremely shocked and upset. I couldnt believe it had happened to me in this country," the woman said after being refused service.

The company responded to the UK-based Jewish newspaper The Jewish Chronicle, saying, "AWG does not refuse to serve people because they support Israel, but do if they support Zionism or genocide."

The UK Lawyers for Israel (UKLFI) group reported the incident to the Equality and Human Rights Commission in the UK. According to the group, the company is violating section 29 of the Equality Act 2020, which prohibits companies from discriminating against potential customers based on protected characteristics, including religion and philosophical belief.

See the rest here:
London plumber rejects Jewish client in antisemitic snub - Ynetnews

An intoxicated plumber who smashed through the Frankies Supermarket glass door at SNPF Plaza was ordered by the Supreme Court to use his skill to assist charitable organisations as part of his community work.

Mani Kopa of Ululoloa and Fatuvalu was recently convicted by the Supreme Court on charges of burglary, intentional damage and armed with a dangerous weapon.

His offending followed a night out at the RSA nightclub and had been drinking Taula Strong. The incident happed around February this year.

According to the summary of facts, the plumber left the RSA bar around 7 pm heading towards the NBS Bank before Police stopped him because he was drunk and told him to take a taxi home.

The defendant left the NBS Building in the taxi and went towards the SNPF Plaza where he stopped the cab got out and went straight to the Frankies Supermarket.

He was armed with a block of cement throwing it at the glass door and later tried to open the door.

Police officers in the area saw what Kopa was doing, stopped him, and arrested him on that night.

The defendant told the Probation Service he had no recollection of the events and his first memory after the drink-up was waking up in the police cell.

Kopa is 25-years-old and is a qualified plumber working for Samoa Water Authority.

The victim of the offending is Frankie Supermarket who said in their Victim Impact Report that no apology or restitution has been paid for the damage caused.

Justice Leiataualesa Daryl Clarke told the defendant that if he had gone into the store and done what was probably on his mind, todays outcome would have been quite different for him.

You are very fortunate that the police were there that evening, said Justice Clarke.

It is clear Mani that when you drink alcohol to the state of drunkenness, you became obscene.

It is then important for you to change your life and your drinking habits. You are therefore to be congratulated for completing the Salvation Army Program.

The Court urged the defendant to continue to learn to control his drinking.

If you return to drinking sessions after work with your friends and drink until you are drunk, there will be a strong likelihood you will come back before the courts, and next time, you might end up at Tanumalala. That would be a huge waste of a very talented young man such as yourself.

Justice Clarke convicted Kopa on all charges. He was sentenced to six months of supervision and ordered 100 hours of community work.

You are a qualified plumber, he said.

Probation Service is urged to utilise your skills to assist charitable organisations and other bodies that may benefit from this community service.

Maiavatele Timothy Fesili was the prosecution and lawyer from the Attorney Generals Office while Quentin Sauaga acted for the defendant.

Go here to read the rest:
Plumber ordered to help charities - Samoa Observer

"If your loved one is a plumber in Minnesota and they now have mesothelioma please make financial compensation a top priority and call attorney Erik Karst of Karst von Oiste anytime at 866-714-6466.

Minnesota Mesothelioma Victims Center

MINNEAPOLIS, MINNESOTA, USA, July 2, 2024 /EINPresswire.com/ -- The Minnesota Mesothelioma Victims Center is urging a journeyman plumber with just diagnosed mesothelioma anywhere in Minnesota or their family to please call attorney Erik Karst of Karst von Oiste at 866-714-6466 to discuss financial compensation. Erik Karst is the managing partner of Karst von Oiste and he is Minnesota's best known mesothelioma attorney. Erik Karst and his colleagues have decades worth of assisting plumbers with mesothelioma and Erik and his team produce remarkable compensation results for their clients.

The group says, "Plumbers who were working before the early 1980s probably had as much exposure to asbestos as a shipyard worker. Many experts consider plumbers to be one of the most at risk work groups for developing mesothelioma and or asbestos exposure lung cancer in the nation. In Minnesota in addition to fixing broken pipes, or water pumps, a plumber would have also worked on residential and commercial boilers and furnaces. Boilers before the 1980s were in most instances covered with asbestos.

"If your loved one is a plumber in Minnesota and they now have mesothelioma please make financial compensation a top priority and call attorney Erik Karst of Karst von Oiste at 866-714-6466. We are certain you will be glad you did." http://www.karstvonoiste.com/

The Minnesota Mesothelioma Victims Center is also appealing to a Navy Veteran or worker who had significant exposure to asbestos before 1983 and who now has lung cancer anywhere in Minnesota to please call them at 866-714-6466 about compensation. A compensation settlement for a person like this might be hundreds of thousands of dollars, and for Veterans there might also be VA Benefits.

The Minnesota Mesothelioma Victims Center is a passionate advocate for people with mesothelioma in Minneapolis, Saint Paul, Rochester, Duluth, Bloomington, Brooklyn Park, Plymouth, Saint Cloud or anywhere else in Minnesota. https://Minnesota.MesotheliomaVictimsCenter.Com

Suggestions from the Mesothelioma Victims Center for people with mesothelioma in Minnesota or nationwide on how to increase potential financial compensation:

* Do you recall the specifics of how you were exposed to asbestos at work, in the military or both-and when this exposure occurred? This is incredibly important information.

* Do you recall the names of coworkers who might have witnessed your exposure to asbestos?

* Did you have more than one job where you might have been exposed to asbestos?

* Do your medical records include a biopsy that confirms the mesothelioma or asbestos exposure lung cancer?"

If a person with mesothelioma or asbestos exposure lung cancer anywhere else in the nation-or their family members would like some very honest suggestions as to what lawyer-law firm to call-please call the Mesothelioma Victims Center anytime at 866-714-6466."We have assembled the most amazing mesothelioma-asbestos exposure lung cancer attorneys in the nation-and we would be honored to make recommendations. We want people like this to receive the best possible compensation results." https://MesotheliomaVictimsCenter.Com

Michael Thomas Minnesota Mesothelioma Victims Center +1 866-714-6466 email us here

You just read:

News Provided By

July 02, 2024, 08:45 GMT

EIN Presswire's priority is source transparency. We do not allow opaque clients, and our editors try to be careful about weeding out false and misleading content. As a user, if you see something we have missed, please do bring it to our attention. Your help is welcome. EIN Presswire, Everyone's Internet News Presswire, tries to define some of the boundaries that are reasonable in today's world. Please see our Editorial Guidelines for more information.

Continue reading here:
Minnesota Mesothelioma Victims Center Urges a Journeyman Plumber with Mesothelioma Anywhere in Minnesota to - EIN News

State Alabama Alaska Arizona Arkansas California Colorado Connecticut Delaware Florida Georgia Hawaii Idaho Illinois Indiana Iowa Kansas Kentucky Louisiana Maine Maryland Massachusetts Michigan Minnesota Mississippi Missouri Montana Nebraska Nevada New Hampshire New Jersey New Mexico New York North Carolina North Dakota Ohio Oklahoma Oregon Pennsylvania Rhode Island South Carolina South Dakota Tennessee Texas Utah Vermont Virginia Washington Washington D.C. West Virginia Wisconsin Wyoming Puerto Rico US Virgin Islands Armed Forces Americas Armed Forces Pacific Armed Forces Europe Northern Mariana Islands Marshall Islands American Samoa Federated States of Micronesia Guam Palau Alberta, Canada British Columbia, Canada Manitoba, Canada New Brunswick, Canada Newfoundland, Canada Nova Scotia, Canada Northwest Territories, Canada Nunavut, Canada Ontario, Canada Prince Edward Island, Canada Quebec, Canada Saskatchewan, Canada Yukon Territory, Canada

Zip Code

Country United States of America US Virgin Islands United States Minor Outlying Islands Canada Mexico, United Mexican States Bahamas, Commonwealth of the Cuba, Republic of Dominican Republic Haiti, Republic of Jamaica Afghanistan Albania, People's Socialist Republic of Algeria, People's Democratic Republic of American Samoa Andorra, Principality of Angola, Republic of Anguilla Antarctica (the territory South of 60 deg S) Antigua and Barbuda Argentina, Argentine Republic Armenia Aruba Australia, Commonwealth of Austria, Republic of Azerbaijan, Republic of Bahrain, Kingdom of Bangladesh, People's Republic of Barbados Belarus Belgium, Kingdom of Belize Benin, People's Republic of Bermuda Bhutan, Kingdom of Bolivia, Republic of Bosnia and Herzegovina Botswana, Republic of Bouvet Island (Bouvetoya) Brazil, Federative Republic of British Indian Ocean Territory (Chagos Archipelago) British Virgin Islands Brunei Darussalam Bulgaria, People's Republic of Burkina Faso Burundi, Republic of Cambodia, Kingdom of Cameroon, United Republic of Cape Verde, Republic of Cayman Islands Central African Republic Chad, Republic of Chile, Republic of China, People's Republic of Christmas Island Cocos (Keeling) Islands Colombia, Republic of Comoros, Union of the Congo, Democratic Republic of Congo, People's Republic of Cook Islands Costa Rica, Republic of Cote D'Ivoire, Ivory Coast, Republic of the Cyprus, Republic of Czech Republic Denmark, Kingdom of Djibouti, Republic of Dominica, Commonwealth of Ecuador, Republic of Egypt, Arab Republic of El Salvador, Republic of Equatorial Guinea, Republic of Eritrea Estonia Ethiopia Faeroe Islands Falkland Islands (Malvinas) Fiji, Republic of the Fiji Islands Finland, Republic of France, French Republic French Guiana French Polynesia French Southern Territories Gabon, Gabonese Republic Gambia, Republic of the Georgia Germany Ghana, Republic of Gibraltar Greece, Hellenic Republic Greenland Grenada Guadaloupe Guam Guatemala, Republic of Guinea, Revolutionary People's Rep'c of Guinea-Bissau, Republic of Guyana, Republic of Heard and McDonald Islands Holy See (Vatican City State) Honduras, Republic of Hong Kong, Special Administrative Region of China Hrvatska (Croatia) Hungary, Hungarian People's Republic Iceland, Republic of India, Republic of Indonesia, Republic of Iran, Islamic Republic of Iraq, Republic of Ireland Israel, State of Italy, Italian Republic Japan Jordan, Hashemite Kingdom of Kazakhstan, Republic of Kenya, Republic of Kiribati, Republic of Korea, Democratic People's Republic of Korea, Republic of Kuwait, State of Kyrgyz Republic Lao People's Democratic Republic Latvia Lebanon, Lebanese Republic Lesotho, Kingdom of Liberia, Republic of Libyan Arab Jamahiriya Liechtenstein, Principality of Lithuania Luxembourg, Grand Duchy of Macao, Special Administrative Region of China Macedonia, the former Yugoslav Republic of Madagascar, Republic of Malawi, Republic of Malaysia Maldives, Republic of Mali, Republic of Malta, Republic of Marshall Islands Martinique Mauritania, Islamic Republic of Mauritius Mayotte Micronesia, Federated States of Moldova, Republic of Monaco, Principality of Mongolia, Mongolian People's Republic Montserrat Morocco, Kingdom of Mozambique, People's Republic of Myanmar Namibia Nauru, Republic of Nepal, Kingdom of Netherlands Antilles Netherlands, Kingdom of the New Caledonia New Zealand Nicaragua, Republic of Niger, Republic of the Nigeria, Federal Republic of Niue, Republic of Norfolk Island Northern Mariana Islands Norway, Kingdom of Oman, Sultanate of Pakistan, Islamic Republic of Palau Palestinian Territory, Occupied Panama, Republic of Papua New Guinea Paraguay, Republic of Peru, Republic of Philippines, Republic of the Pitcairn Island Poland, Polish People's Republic Portugal, Portuguese Republic Puerto Rico Qatar, State of Reunion Romania, Socialist Republic of Russian Federation Rwanda, Rwandese Republic Samoa, Independent State of San Marino, Republic of Sao Tome and Principe, Democratic Republic of Saudi Arabia, Kingdom of Senegal, Republic of Serbia and Montenegro Seychelles, Republic of Sierra Leone, Republic of Singapore, Republic of Slovakia (Slovak Republic) Slovenia Solomon Islands Somalia, Somali Republic South Africa, Republic of South Georgia and the South Sandwich Islands Spain, Spanish State Sri Lanka, Democratic Socialist Republic of St. Helena St. Kitts and Nevis St. Lucia St. Pierre and Miquelon St. Vincent and the Grenadines Sudan, Democratic Republic of the Suriname, Republic of Svalbard & Jan Mayen Islands Swaziland, Kingdom of Sweden, Kingdom of Switzerland, Swiss Confederation Syrian Arab Republic Taiwan, Province of China Tajikistan Tanzania, United Republic of Thailand, Kingdom of Timor-Leste, Democratic Republic of Togo, Togolese Republic Tokelau (Tokelau Islands) Tonga, Kingdom of Trinidad and Tobago, Republic of Tunisia, Republic of Turkey, Republic of Turkmenistan Turks and Caicos Islands Tuvalu Uganda, Republic of Ukraine United Arab Emirates United Kingdom of Great Britain & N. Ireland Uruguay, Eastern Republic of Uzbekistan Vanuatu Venezuela, Bolivarian Republic of Viet Nam, Socialist Republic of Wallis and Futuna Islands Western Sahara Yemen Zambia, Republic of Zimbabwe

Read the original here:
From plumbers to painters: These 10 construction trades employ the most people - The Caledonian-Record