2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH), acetonitrile (high-performance liquid chromatography [HPLC] grade), and the standard syringic acid and vanillic acid were purchased from Sigma-Aldrich (St. Louis, United States). The standard ganoderic acids A, B, C1, and C2 and ganoderenic acid B (≥97%) were purchased from Baoji Chenguang Biotechnology (Baoji, China). The standard protocatechuic acid and gallic acid were purchased from TargetMol (Boston, United States). The standard rutin was purchased from Dr. Ehrenstorfer (Augsburg, Germany). The standard quercetin-3-D -galactoside, coumaric acid, cinnamic acid, quercetin, kaempferol, and hydroxybenzoic acid were purchased from Shanghai YuanYe Biotechnology (Shanghai, China). pBR322 plasmid DNA was purchased from Sangon Biotech. (Shanghai, China). RNA Isolation kit, TransScript All-in-One First-Strand cDNA Synthesis SuperMix kit, and other qPCR reagents were purchased from TransGen Biotech (Beijing, China). Ascorbic acid, pyrogallol, hydrogen peroxide, and all other chemicals and solvents were purchased from Shanghai Chemical Reagent Company (Shanghai, China).
Rna isolation kit
Manufactured by Transgene
Sourced in China
The RNA Isolation Kit is a laboratory product designed to extract and purify ribonucleic acid (RNA) from various biological samples. It utilizes a proprietary method to effectively isolate and concentrate RNA for downstream applications.
Automatically generated - may contain errors
Lab products found in correlation
Cinnamic acid, by Yuanye Bio-Technology (1 mentions)
Coumaric acid, by Yuanye Bio-Technology (1 mentions)
Transscript all in one first strand cdna synthesis supermix kit, by Transgene (1 mentions)
Vanillic acid, by Merck Group (1 mentions)
2 2 azino bis, by Merck Group (1 mentions)
Kaempferol, by Yuanye Bio-Technology (1 mentions)
Quercetin, by Yuanye Bio-Technology (1 mentions)
Ascorbic acid, by Shanghai Chemical Reagent (1 mentions)
Hydrogen peroxide, by Shanghai Chemical Reagent (1 mentions)
Cdna transcription kit, by Thermo Fisher Scientific (1 mentions)
Lightcycler 480, by Roche (1 mentions)
Superscript 2 reverse transcriptase, by Thermo Fisher Scientific (1 mentions)
Truseq rna sample preparation kit v2, by Illumina (1 mentions)
Hiseq 4000 platform, by Illumina (1 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions)
4 protocols using rna isolation kit
1
Antioxidant Compounds and Reagents Protocol
2
Bone Tissue RNA Extraction and RT-PCR
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The bone tissues which weigh 100 mg–200 mg were ground into a powder with liquid nitrogen. Total RNA was isolated from bone tissue using an RNA isolation kit (Transgene, China) following the manufacturer's instructions. The total RNA which weigh 2 μg was reverse-transcribed by a cDNA transcription kit (Thermo Fisher Scientific, USA). RT-PCR was executed by using SYBR Green I Master on LightCycler®480 (ROCHE, USA) instrument with primers (RT-PCR detection primer sequence was shown in Table 2 ). 18S rRNA was used as a normalized control. The mRNA levels of target genes were calculated with the ΔΔct method compared to normalized control.
3
Transcriptome Analysis of Floral Buds
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Total RNA of both LD6A and LD6B lines were extracted from the floral bud (tetrad stage) using an RNA Isolation Kit (TransGen Biotech, Beijing, China). From each sample, 3 µg of the total RNA (RIN ≥8) was used for transcriptome cDNA library construction with a TruSeq™ RNA Sample Preparation Kit v2 (Illumina, San Diego, CA, USA). RNA purification beads with oligo (dT) were used to separate mRNA from the total RNA. After breaking the mRNA into short fragments in fragmentation buffer, double-stranded cDNA was synthesized (Super ScriptII reverse transcriptase, Invitrogen, Carlsbad, CA, USA) and purified (Agencourt AMPure XP-Medium, Agencourt, Carlsbad, CA, USA). The short cDNA fragments were end-repaired with an A-tail addition and connected with adapters. After agarose gel electrophoresis, suitable fragments were used as templates for PCR amplification. The Agilent 2100 Bioanalyzer (Agilent, Santa Clara, CA, USA) and an ABI StepOnePlus Real-Time PCR System were respectively used for quantification and qualification of the sample cDNA library. Solexa sequencing was performed by BGI (Shenzhen, China) using an Illumina HiSeq 4000 platform. The raw sequencing files of these six samples (FASTQ files) are accessible from the NCBI Sequence Read Archive (SRA) database under Accession Number PRJNA577562.
4
Comparative Cardiac Fibroblast Transcriptomics
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To compare gene expressing profiles of different cardiac fibroblasts, we analyzed MICFs with the CPM data of Sham P1 fibroblasts and Sham P56 fibroblasts from GSE95755 (Quaife-Ryan et al., 2017 (link)). The read count data of MICFs was transformed into CPM data for fold change analysis. Hierarchical clustering was performed with 2,585 genes which log2 (fold change) > 2 and GO enrichment analysis was performed using the OmicShare tools1 .
After a 4 week induction, Myh6-mCherry+ iCMs from GMTMS- and GMTMM-transduced MICFs were collected by FACS (BD Aria3). RNA was extracted using an RNA Isolation Kit (TransGene, ET111). RNA was sequenced by PE150. For data analysis, data processing was performed with MatLab 2019a. FPKM was normalized with the “quantilenorm” function over the 3 samples and transferred as log2 (FPKM+1). Hierarchical clustering of genes and samples was performed after genes that varied less than 1 through the 3 samples were filtered out. GO enrichment was analyzed with the websitehttp://geneontology.org/ . Principal component analysis (PCA) was performed using RNA sequencing data of GSE95755 including fibroblasts and cardiomyotytes from both neonatal and adult mice (Quaife-Ryan et al., 2017 (link)). All data is normalized and top 1,000 differentially expressed genes enriched for GO analysis above are used of PCA input.
After a 4 week induction, Myh6-mCherry+ iCMs from GMTMS- and GMTMM-transduced MICFs were collected by FACS (BD Aria3). RNA was extracted using an RNA Isolation Kit (TransGene, ET111). RNA was sequenced by PE150. For data analysis, data processing was performed with MatLab 2019a. FPKM was normalized with the “quantilenorm” function over the 3 samples and transferred as log2 (FPKM+1). Hierarchical clustering of genes and samples was performed after genes that varied less than 1 through the 3 samples were filtered out. GO enrichment was analyzed with the website
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