Cell culture reagents were obtained from Invitrogen (Carlsbad, CA, USA). RNA extraction reagents were purchased from Qiagen (Valencia, CA, USA). Reagents for cDNA synthesis and qRT-PCR were from Bio-Rad (Hercules, CA, USA). Murine M-CSF, IFN-γ, IL-4, and IL-10 were obtained from PeproTech (Rocky Hill, NJ, USA). Azoxymethane and LPS were obtained from Sigma-Aldrich (St. Louis, MO, USA). The DSS used was purchased from TdB Consultancy (Uppsala, Sweden). 14C L-Arg was obtained from Perkin-Elmer (Waltham, MA, USA).
Rna extraction reagent
Manufactured by Qiagen
Sourced in United States, Netherlands
RNA extraction reagents are laboratory products designed to efficiently and reliably isolate and purify RNA from a variety of biological samples. These reagents provide the necessary chemical components and protocols to enable the extraction and separation of RNA molecules from cells, tissues, or other sources, preserving their integrity for downstream applications such as analysis or experimentation.
Automatically generated - may contain errors
Lab products found in correlation
Azoxymethane, by Merck Group (2 mentions)
Interleukin 4 (il 4), by Thermo Fisher Scientific (2 mentions)
Ifn γ, by Thermo Fisher Scientific (2 mentions)
Reagents for cdna synthesis and qrt pcr, by Bio-Rad (2 mentions)
14c l arg, by PerkinElmer (2 mentions)
Murine m csf, by Thermo Fisher Scientific (2 mentions)
Cell culture reagents, by Thermo Fisher Scientific (2 mentions)
Il 10, by Thermo Fisher Scientific (2 mentions)
Lipopolysaccharides (lps), by Merck Group (2 mentions)
Nanodrop 2000c, by Thermo Fisher Scientific (2 mentions)
Mas5 algorithm, by Thermo Fisher Scientific (1 mentions)
Recombinant macrophage colony stimulating factor m csf, by Thermo Fisher Scientific (1 mentions)
Bca protein assay, by Thermo Fisher Scientific (1 mentions)
Truseq stranded messenger rna library preparation kit, by Illumina (1 mentions)
Nextseq 500 system, by Illumina (1 mentions)
Sybr premix ex taq, by Takara Bio (1 mentions)
Real time pcr system, by Thermo Fisher Scientific (1 mentions)
Primescripttm 2 1st strand cdna synthesis kit, by Takara Bio (1 mentions)
Sybr premix ex taqtm 2, by Takara Bio (1 mentions)
Rnase free dnase set, by Qiagen (1 mentions)
8 protocols using rna extraction reagent
1
Murine Inflammatory Cytokine Assay
2
Transcriptional Profiling of Gastric Cancer
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Primary gastric tumors were obtained from National Cancer Centre, Singapore with approvals from Research Ethics Review Committee, and signed patient informed consent. Total RNA was extracted using Qiagen RNA extraction reagents (Qiagen, Venlo, Limburg, Netherlands) according to the instructions of the manufacturer and hybridized to Affymetrix Human Genome U133 plus Genechips (HG-U133 Plus 2.0, Affymetrix, Santa Clara, CA, USA). Raw data obtained after chip-scanning was pre-processed using the MAS5 algorithm (Affymetrix). Data were subjected to Log 10 transformation followed by median-centered across all probe sets for each sample (primary tumor). The centering is such that the median of expression in each sample is zero. 160 were classified by pathological diagnosis into diffuse-type GCs (68 cases) and intestinal-type (92 cases).
Formalin-fixed and paraffin-embedded (FFPE) primary tumor and matched normal tissue samples from 118 patients with gastric cancer were obtained from the Department of Pathology at the National University Hospital System, Singapore, under an institutionally approved protocol.
Formalin-fixed and paraffin-embedded (FFPE) primary tumor and matched normal tissue samples from 118 patients with gastric cancer were obtained from the Department of Pathology at the National University Hospital System, Singapore, under an institutionally approved protocol.
3
Murine Inflammatory Cytokine Assay
4
Macrophage Differentiation and Characterization
5
Profiling RNA Differences in CAFs vs NAFs
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Total RNA was extracted from a pair of CAFs and NAFs with RNA extraction reagents (Qiagen, Valencia, CA, USA). RNA libraries were prepared with a TruSeq stranded messenger RNA library preparation kit (Illumina), and RNA sequencing was performed with an Illumina NextSeq 500 system following protocols for 1 9 75 bp sequencing provided. Reads for each sample were mapped to the reference genome (human hg19) by TopHat (version 2.0.13). The aligned results were added to Cuffdiff (version 2.2.0) to find differentially expressed genes. To estimate the functions of the differentially expressed RNAs in CAFs compared with NAFs, we performed Gene Ontology analysis using the DAVID web application (https://david.ncifcrf.gov).
6
Polycyclic Aromatic Hydrocarbon Analysis
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Chr,
B[a]P, and dibenzo[a,h]anthracene (DB[a,h]A) were supplied
by Wako Pure Chemical Industries, Ltd.
(Osaka, Japan); fluoranthene (FA) was supplied by Tokyo Kasei Kogyo
Co., Ltd. (Tokyo, Japan); anthracene (Ant) and Py were supplied by
Nacalai Tesque, Inc. (Kyoto, Japan); benzo[c]fluorene
(B[c]Fl) was supplied by Dr. Ehrenstorfer GmbH (Augsburg,
Germany); dibenzo[a,h]pyrene (DB[a,h]P), DB[a,i]P, and DB[a,l]P were supplied by AccuStandard, Inc. (New Haven,
CT,
USA); other test chemicals were supplied by Sigma-Aldrich Co. LLC.
(St. Louis, MS, USA). The purities of many test chemicals were 99–100%,
those of FA, B[c]Fl, B[b]FA, and
benzo[ghi]perylene (BPe) were ≥98%, and that
of DB[a,h]A was 96%. The mite antigen,
Df body extract, was supplied by ITEA Inc. (Tokyo, Japan). Dulbecco’s
modified Eagle’s medium (DMEM) and fetal bovine serum (FBS)
were Gibco products purchased from Thermo Fisher Scientific (Waltham,
MA, USA). The RNA extraction reagent was supplied by Qiagen N. V.
(Venlo, the Netherlands). Dimethyl sulfoxide (DMSO) was supplied by
Wako Pure Chemical Industries, Ltd.
B[a]P, and dibenzo[a,h]anthracene (DB[a,h]A) were supplied
by Wako Pure Chemical Industries, Ltd.
(Osaka, Japan); fluoranthene (FA) was supplied by Tokyo Kasei Kogyo
Co., Ltd. (Tokyo, Japan); anthracene (Ant) and Py were supplied by
Nacalai Tesque, Inc. (Kyoto, Japan); benzo[c]fluorene
(B[c]Fl) was supplied by Dr. Ehrenstorfer GmbH (Augsburg,
Germany); dibenzo[a,h]pyrene (DB[a,h]P), DB[a,i]P, and DB[a,l]P were supplied by AccuStandard, Inc. (New Haven,
CT,
USA); other test chemicals were supplied by Sigma-Aldrich Co. LLC.
(St. Louis, MS, USA). The purities of many test chemicals were 99–100%,
those of FA, B[c]Fl, B[b]FA, and
benzo[ghi]perylene (BPe) were ≥98%, and that
of DB[a,h]A was 96%. The mite antigen,
Df body extract, was supplied by ITEA Inc. (Tokyo, Japan). Dulbecco’s
modified Eagle’s medium (DMEM) and fetal bovine serum (FBS)
were Gibco products purchased from Thermo Fisher Scientific (Waltham,
MA, USA). The RNA extraction reagent was supplied by Qiagen N. V.
(Venlo, the Netherlands). Dimethyl sulfoxide (DMSO) was supplied by
Wako Pure Chemical Industries, Ltd.
7
BIRC3 Expression Analysis in Injured Nerves
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Total RNA from injured sciatic nerves was isolated using RNA extraction reagent (Qiagen, Valencia, CA, USA) to determine the BIRC3 expression level. RNA samples were reverse transcribed to cDNA using a cDNA Transcription Kit (Qiagen) following the manufacturer’s instructions (Applied Biosystems® 2720, Foster City, CA, USA). According to the manufacturer’s protocol, we performed quantitative real-time polymerase chain reaction (qRT-PCR) with SYBR Premix Ex Taq (TaKaRa, Shanghai, China) using a polymerase chain reaction (PCR) system (Applied Biosystems® Real-Time PCR System). The forward and reverse PCR primers are listed in Table 3
8
Quantitative Gene Expression Analysis
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RNA was isolated and extracted using the RNA extraction reagent (QIAGEN, Germany) according to the manufacturer’s protocol. The genomic DNA (gDNA) was cleaved using the RNAse-Free DNase Set from QIAGEN, a company based in Germany. The RNA samples were assessed for purity using the NanoDrop 2000c (Thermo Fisher Scientific, United States) and gel electrophoresis. The reverse transcription PCR was performed using the PrimeScriptTM II 1st Strand cDNA Synthesis Kit from TaKaRa, a company based in China. The SYBR Premix Ex TaqTM II, manufactured by TaKaRa in China, was employed for quantitative reverse transcription polymerase chain reaction (qRT-PCR) on a 7,500 series real-time fluorescence quantitative cycler manufactured by Bio-RAD in the United States of America. The primers for the qRT-PCR test were prepared using the Primer Premier 5.0 software. The ACTIN gene served as the reference gene (Song et al., 2021 (link)). The primers utilized for qRT-PCR analysis are listed in Supplementary Table S2 . Each experiment was repeated three times in triplicate, and a total of three biological replicates were undertaken. The gene expression levels were determined using the 2-△△CT method.
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