Total RNA from sorted cells was extracted using Aurum Total RNA kit (Bio-Rad Laboratories, Inc.). RNA integrity score was determined by Agilent 2100 Bioanalyzer using the Agilent RNA 6000 Pico kit (Agilent Technologies). Samples were prepared for Illumina sequencing using NEB’s Ultra Directional RNA Library Prep Kit for Illumina (NEB#7420). Libraries were sequenced with a 50 bp SR run on Illumina HiSeq 2500 using a V3 flow cell. Sequenced reads were compared to available murine Ensembl 70 genes using mouse genome build (GRCm38), and expression was compared between macrophage subtypes using two separate analysis pipelines: RSEM/EdgeR and topaht2/cuffdiff. Depending on the pipeline, between ~1500–2500 genes were found to be differentially expressed (FDR < 0.05), with a wide overlap in results between the two pipelines. Significance values presented were from the topaht2/cuffdiff analysis. Gene ontology enrichment analysis was performed with the DAVID Bioinformatics Resources 6.7 software.
Ultra directional rna library prep kit for
Manufactured by Illumina
The Ultra Directional RNA Library Prep Kit for Illumina is a laboratory equipment product designed for the preparation of directional RNA libraries for sequencing on Illumina platforms. The kit provides the necessary reagents and protocols to generate high-quality directional RNA libraries from total RNA samples.
Automatically generated - may contain errors
Lab products found in correlation
Hiseq 2500, by Illumina (4 mentions)
Aurum total rna kit, by Bio-Rad (2 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (2 mentions)
Agilent rna 6000 pico kit, by Agilent Technologies (2 mentions)
Turbo dna free dnase kit, by Thermo Fisher Scientific (2 mentions)
Ribo zero human and bacterial rrna removal kits, by Illumina (2 mentions)
Nebnext poly a mrna magnetic isolation module, by Illumina (2 mentions)
Trizol solution, by Roche (2 mentions)
Ribo zero plus rrna depletion kit, by Illumina (2 mentions)
Rneasy mini kit, by Qiagen (2 mentions)
Rnase free dnase set, by Qiagen (2 mentions)
Novaseq 6000, by Illumina (1 mentions)
Hiseq 4000 platform, by Illumina (1 mentions)
Dnase 1, by Qiagen (1 mentions)
Agilent bioanalyzer 2100 system, by Agilent Technologies (1 mentions)
Fastrna pro blue kit, by MP Biomedicals (1 mentions)
Novaseq 6000 next generation sequencer, by Illumina (1 mentions)
10 protocols using ultra directional rna library prep kit for
1
RNA-seq Analysis of Macrophage Subtypes
2
Campylobacter jejuni RNA Extraction from Diarrhea
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Campylobacter jejuni strain CG8421 infected diarrhea was weighed and added 1:1 in RNA Later reagent as quickly as possible after it was produced and frozen at −80°C. Samples came from volunteers who were not receiving antibiotics and therefore represent untreated infections. Preserved samples were thawed on ice and total RNA extracted via Trizol-chloroform phase separation. DNA was removed (Turbo DNA-free DNAse kit, Ambion), rRNA depleted (Ribo-Zero human and bacterial rRNA removal kits, Illumina), and libraries built for Illumina Sequencing (Ultra Directional RNA Library Prep Kit for Illumina, New England Biolabs).
3
Transcriptome Analysis by RNA-Seq
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was extracted using TRIzol solution (Roche) according to the manufacturer’s instructions. cDNA libraries were prepared from 500 ng of each total RNA sample for massive parallel sequencing using the NEBNext Poly(A) mRNA Magnetic Isolation Module and an Ultra Directional RNA Library Prep Kit for Illumina (New England BioLabs). The cDNA library contained DNA ranging from 400 to 1000 bp, including the adaptor sequences. RNA-seq was performed on an Illumina HiSeq 2500 using 50-nucleotide read length single-end sequencing at Macrogen. The sequence reads were mapped to the human genome (hg19), and the expression values for genes were calculated as reads per kilobase of exon per million mapped reads (RPKM) using the Biowardrobe platform. Heatmaps were generated using the Morpheus software developed by the Broad Institute.
4
Transcriptome Analysis of E. coli Growth Phases
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The E. coli cells were cultured in 5 ML of the fresh media and rotated at 200 rpm at 37 ℃. Independent cultures as biological replicates of transcriptomes were performed. The cell cultures of both exponential and stationary growth phases were collected, as described previously [49] , [50] . The total RNAs were purified using RNeasy Mini Kit (QIAGEN) and RNase-Free DNase Set (QIAGEN) according to the product instructions. The eluted RNAs were suspended with RNase-free water and subsequently subjected to RNAseq. The rRNAs were removed using Ribo-Zero Plus rRNA Depletion Kit (Illumina), and mRNA preparation was performed with Ultra Directional RNA Library Prep Kit for Illumina (NEBNext). The paired-end sequencing (150 bp × 2) was performed with Novaseq6000 (Illumina) by Chemical Dojin Co. Ltd.
5
Directional RNA-seq Library Prep
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was extracted using TRIzol solution (Roche) according to the manufacturer's instructions. The cDNA library was prepared from 500 ng of each total RNA sample for massive parallel sequencing using an NEBNext Poly(A) mRNA Magnetic Isolation Module and an Ultra Directional RNA Library Prep Kit for Illumina (NEB). The cDNA library produced ranged from 400 to 1000 bp, including the adaptor sequences.
6
Campylobacter jejuni RNA Extraction from Diarrhea
7
Whole-Transcriptome Analysis of Mycobacterium tuberculosis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was isolated from Mtb strains using FastPrep-24 with a FastRNA Pro Blue Kit (MP Biomedicals) following the manufacturer’s instructions. Total RNA fragmentation was performed using the ultrasonic method (140–160 bp) (Covaris M220) after DNase I (QIAGEN) treatment and ribosomal RNA removal (Epicentre). Random primers were used to synthesize the first and second strands, and dTTP was replaced with dUTP for complementary (cDNA) synthesis. Ultra-™Directional RNA Library Prep Kit for Illumina (NEB) was used according to the manufacturer’s instructions. The final library products were purified using 0.8× beads (Beckman) and assessed using an Agilent Bioanalyzer 2100 system (Agilent). The Illumina HiSeq 4000 platform was used for the whole-transcriptome analysis.
8
Transcriptome Profiling of E. coli
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The E. coli cells were collected at the exponential growth phase (i.e., 5 × 107 – 2×108 cells/ml) and subjected to RNA sequencing, as described previously (Matsui et al., 2023 (link); Liu et al., 2020 (link)). In brief, the bacterial growth was stopped by mixing with the iced 10% phenol ethanol solution. The cell pellet was collected to purify the total RNAs using the RNeasy Mini Kit (QIAGEN) and RNase-Free DNase Set (QIAGEN) according to product instructions. The paired-end sequencing (150 bp ×2) was performed using the Novaseq6000 next-generation sequencer (Illumina). The rRNAs were removed from the total RNAs using the Ribo-Zero Plus rRNA Depletion Kit (Illumina), and the mRNA libraries were prepared using the Ultra Directional RNA Library Prep Kit for Illumina (NEBNext). Biological replicates were performed for all conditions (N = 2–4). The raw datasets were deposited in the DDBJ Sequence Read Archive under the accession number DRA013662.
9
RNA Extraction and Sequencing of Sorted Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
RNA extraction was performed as previously described (17 (link)). Briefly, all RNA species from sorted cells were extracted using the Aurum Total RNA kit (Bio-Rad Laboratories, Inc.). RNA integrity was scored by Agilent 2100 Bioanalyzer using the Agilent RNA 6000 Pico kit (Agilent Technologies). Samples were prepared for Illumina sequencing using NEB's Ultra Directional RNA Library Prep Kit for Illumina (NEB#7420). Libraries were sequenced with a 50 bp SR run on Illumina HiSeq 2500 using a V3 flow cell.
10
Illumina RNA Sequencing Protocol
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!