Zymosan was purchased from Sigma-Aldrich (St. Louis, MO, USA). The antibodies used for Western blotting were as follows: anti-phospho STAT6 (Tyr-641), anti-STAT6, anti-PPARγ, anti-CD36, anti-MMR, and anti-arginase-1 (Arg1) from Cayman Chemical Co (Ann Arbor, MI, USA), anti-phospho JAK3 (Tyr-980/981), anti-JAK3 (Cell signalling Technology, Danvers, MA), and anti-β-actin from Sigma-Aldrich. Pierce BCA protein assay kit was purchased from Thermo Scientific (Rockford, IL, USA). The gene-specific relative RT-PCR kit was obtained from Invitrogen Life Technologies (Carlsbad, CA, USA). M-MLV reverse transcriptase was from Enzynomics (Seoul, Korea). DNA polymerase Klenow fragment and dNTPs were obtained from Intron Biotechnology (Seoul, Korea).
Dntps
Manufactured by iNtRON Biotechnology
DNTPs are the building blocks of DNA. They are essential for DNA synthesis and amplification processes in various molecular biology applications.
Automatically generated - may contain errors
Lab products found in correlation
Zymosan, by Merck Group (2 mentions)
Gene specific relative rt pcr kit, by Thermo Fisher Scientific (2 mentions)
Anti β actin, by Merck Group (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
M mlv reverse transcriptase, by Enzynomics (2 mentions)
Anti pparγ, by Cayman Chemical (1 mentions)
Anti cd36, by Cayman Chemical (1 mentions)
Anti mmr, by Cayman Chemical (1 mentions)
Taq polymerase, by iNtRON Biotechnology (1 mentions)
Anti stat6, by Cell Signaling Technology (1 mentions)
Anti cd36, by Cell Signaling Technology (1 mentions)
Anti phospho stat6 tyr 641, by Cell Signaling Technology (1 mentions)
Anti pparγ, by Cell Signaling Technology (1 mentions)
Anti anxa1, by Cell Signaling Technology (1 mentions)
Anti arg1, by Cell Signaling Technology (1 mentions)
As1517499, by Axon Medchem (1 mentions)
Nucleospin gel and pcr clean up kit, by Macherey-Nagel (1 mentions)
Agencourt ampure xp, by Beckman Coulter (1 mentions)
Nucleospin soil kit, by Macherey-Nagel (1 mentions)
I taq, by iNtRON Biotechnology (1 mentions)
4 protocols using dntps
1
Zymosan-Induced Macrophage Activation
2
PCR Profiling of Pseudomonas aeruginosa Virulence
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The PCR was carried out by using primers targeting (oprI, toxA and lasB) virulence genes [10] ., the PCR assay included 25 μl final reaction mixture which consisted 5 μl of (taq polymerase, reaction buffer., MgCl and DNTPs) (Intron biotechnology, Korea) and 2 μl of each 10 P mol forward and reverse primers specific for these virulence genes, 5 μl from DNA template and 13 μl from (ddH 2 O) to complete the volume to 25 μl final reaction mixture .The reaction tubes were cycled in thermal cycler machine (Techne, England). The PCR program was performed with an initial denaturation for 5 min at 94°C, then 35 cycles of denaturation for 1min at 94°C, annealing for 1min at 58°C and 60°C for lasB gene, extension for 1 min at 72°C and final extension for 5 min at 72°C. The amplified products were resolved in %2 agarose gel electrophoresis stained with ethidium bromide and visualized under UV light [10] . Primers used for amplification of virulence genes of P. aeruginosa isolates as in Table-1.
3
Investigating Anti-Inflammatory Mechanisms
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The following reagents were used throughout this study: zymosan (Sigma-Aldrich, St. Louis, MO, USA), AS1517499 (Axon Medchem BV, Groningen, Netherlands), and recombinant mouse AnxA1 (ab202184; Abcam, Cambridge, UK). For ELISA, we used the mouse TSG-6 ELISA kit (RayBiotech Life, Peachtree Corners, GA, USA) and mouse AnxA1 ELSIA kit (ab264613; Abcam, Boston, MA, USA). The antibodies used for Western blotting were as follows: anti-AnxA1, anti-phospho STAT6 (Tyr-641), anti-STAT6, anti-PPARγ, anti-CD36, anti-MMR, and anti-Arg1 from Cell Signaling Technology (Danvers, MA, USA) and anti-β-actin from Sigma-Aldrich. The Pierce BCA protein assay kit was purchased from Thermo Fisher Scientific (Rockford, IL, USA). The gene-specific relative RT-PCR kit was obtained from Invitrogen Life Technologies (Carlsbad, CA, USA). M-MLV reverse transcriptase was obtained from Enzynomics (Seoul, Korea). The Klenow fragment of DNA polymerase and dNTPs were obtained from Intron Biotechnology (Seoul, Korea).
4
Soil Microbial DNA Extraction and Sequencing
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DNA was extracted from soil microcosms using a modified protocol for the NucleoSpin Soil kit (Macherey-Nagel) (Houghton and Stewart, 2019 (link)). DNA was amplified using universal primers for the V4 region (515F, 806R) of the 16S rRNA gene (Caporaso et al., 2011 (link)). PCR was carried out in 50 μL reaction volumes containing 100 μM dNTPs, 0.5 μM primers, 1 U i-Taq (iNtRON Biotechnology) and 7 μL of an enhancer solution (2.7 M betaine, 0.2 M trehalose, 6.7 mM DTT, 0.06 mg ml−1 BSA and 0.07% DMSO). The final concentration of MgCl2 was 1.5 mM. DNA templates from microcosms were used at final concentrations of 10–50 ng reaction−1. Three PCR amplicons (~300 bp) for each sample were pooled. The amplicons were then purified using the NucleoSpin Gel and PCR Clean-up kit (Macherey-Nagel) and Agencourt AmPure XP (Beckman Coulter). Amplicon libraries using the PCR products were prepared and sequenced by Macrogen Inc. Sequencing data was deposited in the NCBI BioProject database (PRJNA766707 and PRJNA546003).
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