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Molecular biology reagents

Manufactured by Thermo Fisher Scientific
Sourced in United States, Germany, United Kingdom, Lithuania

Molecular biology reagents are a class of laboratory supplies used in various molecular biology techniques. These reagents provide the necessary components for DNA and RNA analysis, including buffers, enzymes, dyes, and other essential materials required for procedures such as PCR, DNA sequencing, and gel electrophoresis. The core function of these reagents is to facilitate the manipulation and study of genetic material in a controlled laboratory environment.

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19 protocols using molecular biology reagents

1

Reagents for Cell and Molecular Biology

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Cell culture reagents, molecular biology reagents and kits were purchased from Thermo Fisher Scientific and all other chemicals from Sigma, unless stated otherwise.
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2

Characterization of Enzyme Activities

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Unless otherwise stated, chemicals employed were of the highest grade obtainable. 2‐Chlorobenzoates, 3‐chlorobenzoates, 4‐chlorobenzoates, dichlorodiphenyltrichloroethane (DDT), endosulfan and permethrin were purchased from Merck Millipore, Burlington, MA, USA. 1‐Chloro‐2,4 dinitrobenzene (CDNB), ethacrynic acid, hydrogen peroxide, cumene hydroperoxide, GSH, NADPH and glutathione reductase were all purchased from Sigma‐Aldrich (St Louis, MO, USA). Molecular biology reagents were purchased from Thermofisher Scientific, Waltham, MA, USA. QuickChange lightning site‐directed mutagenesis kit was purchased from Agilent Technologies (Santa Clara, CA, USA).
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3

Detailed Microbiome Profiling Protocol

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All media and chemicals were purchased from Oxoid (Basingstoke, Hampshire, United Kingdom) and Sigma-Aldrich (St. Louis, MO, United States), respectively, except where differently indicated. Anaerobic systems and molecular biology reagents were purchased from Thermo Fisher Scientific (Waltham, MA, United States). Oligonucleotide and Sanger sequencing services were provided by Bio-Fab Research (Rome, Italy).
The type strains used in this study are listed in Table 1. All the strains were purchased from DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Deutschland) and cultivated according to DSMZ culture condition specifications.
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4

Monoclonal Antibody Generation for RC

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RC and its γδ subunit were isolated as previously described [40 (link),51 (link)]. The monoclonal antibodies (mAbs) against RC, among them IIIG5 from mice and IC3 from rats, were generated and isolated as previously described [40 (link)]. The murine mAbs against the human α2A domain, JA202 and JA218, were a generous gift from D. Tuckwell (formerly of the University of Manchester, United Kingdom) [40 (link),42 (link)]. PCR primers were obtained from Eurofins (Eurofins Genomics, Germany) and are written in 5′-3′ direction. Restriction enzymes and molecular biology reagents were from Thermo Fisher Scientific (Germany) unless otherwise stated. Cloning products and expression vectors were validated by DNA sequencing (Eurofins Genomics).
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5

Preparation of Culture Media Reagents

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For the preparation of culture media, the reagents used were obtained from the following companies: Reachim (Moscow, Russia), AppliChem (Darmstadt, Germany) and Panreac (Barcelona, Spain). Biochemical reagents were obtained from Sigma-Aldrich (Sigma, St. Louis, MO, USA), Serva (Heidelberg, Germany) and Fluka (Seelze, Germany), and molecular biology reagents were obtained from Thermo Fischer Scientific (Waltham, MA, USA), Zymo Research (Irvine, CA, USA), New England BioLabs (Hitchin, UK) and Biocom (Moscow, Russia).
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6

Yeast Growth Assay in Glycerol-Ethanol Media

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Yeast strains were cultured in YP (1% yeast extract, 2% peptone) or synthetic complete (0.67% yeast nitrogen base without amino acids, SC-Ura drop-out mix) medium with 2% glycerol and 2% ethanol as carbon sources unless otherwise mentioned69 . Both media were supplemented with 2% agar for preparing solid media. Yeast was grown in 30 C with 230 rpm shaking in 20 mL volume in 100 mL flasks or in 50 mL volume in 250 mL flasks. Growth curves were recorded with Bioscreen C incubator (Oy Growth Curves Ab, Helsinki, Finland) as described before21 (link).
Chemicals were obtained from Merck KGaA (Darmstadt, Germany) and molecular biology reagents from Thermo Fisher Scientific (Waltham, MA, USA) unless otherwise mentioned. Lactose was obtained from VWR (Helsinki, Finland), and l-rhamnose and l-sorbose were obtained from Fluka (Charlotte, NC, USA). Cellotriose and cellotetraose were obtained from Carbosynth (Compton, Berkshire, UK) and α -sophorose from SERVA Electrophoresis GmbH (Heidelberg, Germany).
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7

Synthesis and Characterization of AMPS-HDB

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NAD+ (crystallised lithium salt, ca. 100%), NADH (disodium salt, grade II, 98%), ADP (disodium salt, 98%) and crystalline bovine serum albumin (fraction V) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Molecular biology reagents were obtained from Invitrogen (Thermo Fisher Scientific, Waltham, MA, USA). The nucleotide analogue AMPS-HDB was synthesized as previously described by Vollmer et al., 1994 [38 (link)] for the 5′-O-[S-(4-bromo-2,3-dioxobutyl)]-thiophosphate. The replacement of the 4-bromo group by hydroxyl-group was achieved by incubation (5 h) in 0.03 M 2-(N-morpholino)ethanesulfonic acid (MES) buffer, pH 7.0 at 25 °C as previously described [38 (link)]. The structure was verified by electrospray tandem mass spectrometry (ESI/MS/MS) as described in [51 (link)].
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8

Molecular Reagents for Cell Culture

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All tissue culture reagents were purchased from Life Technologies (Paisley, UK); unless otherwise mentioned, other reagents were obtained from Sigma Aldrich (Poole, Dorset, UK). Molecular biology reagents were purchased from Invitrogen (Paisley, UK) and Qiagen Ltd (Crawley,UK). Primers for RT-PCR and qPCR were from MWG Biotech (Ebersberg, Germany) and Qiagen, respectively. The NPP1 antibody was obtained from Thermo Fisher Scientific Inc. (Illinois, USA) and the β-actin antibody from Abcam (Cambridge, UK)
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9

Cell Culture Reagents and Molecules

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Tissue culture reagents were obtained from Corning Life Sciences B.V. (The Netherlands). Alpha-modified minimal essential media (α-MEM) without l-glutamine was purchased from Lonza (UK) and fetal bovine serum (FBS) was from Biosera (UK). Molecular biology reagents were purchased from Invitrogen (UK). Dexamethasone, alkaline phosphatase kits, and all other biochemical reagents were of analytical grade from Sigma (UK) unless otherwise stated.
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10

Molecular Reagents Utilized in Cell Culture

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All tissue culture reagents were purchased from Life Technologies (Paisley, UK); unless otherwise mentioned, other reagents were obtained from Sigma–Aldrich (Poole, Dorset, UK). Molecular biology reagents were purchased from Invitrogen (Paisley, UK) and Qiagen Ltd (Crawley, UK). Primers for RT‐PCR and qPCR were from MWG Biotech (Ebersberg, Germany) and Qiagen, respectively. The NPP1 antibody was obtained from Thermo Fisher Scientific (Loughborough, UK) and the β‐actin antibody from Abcam (Cambridge, UK).
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