Truseq stranded total rna with ribo zero gold sample preparation kit

Manufactured by Illumina

Sourced in United States

The TruSeq Stranded Total RNA with Ribo-Zero Gold Sample Preparation Kit is a laboratory equipment product designed for the preparation of RNA samples for sequencing. The kit provides a method to remove ribosomal RNA from total RNA, enabling the enrichment and sequencing of non-coding RNA species.

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11 protocols using truseq stranded total rna with ribo zero gold sample preparation kit

Comprehensive RNA Extraction and Sequencing

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RNA extractions were done on blood using miRNeasy Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. The quantity and quality of isolated RNA were determined using the NanoDrop 2000 Spectrophotometer (Thermo Fischer Scientific, Waltham, USA), Qubit^® RNA HS Assay Kit (Invitrogen, Waltham, USA), and 2200 TapeStation Automated Electrophoresis System (Agilent Technologies, Santa Clara, USA). The samples had RNA integrity numbers between 2.2 and 8.8.
Ribosomal RNA was removed, and sequencing libraries were prepared using the TruSeq Stranded Total RNA Sample Preparation Kit with Ribo-Zero Gold (Illumina, San Diego, USA), following the manufacturer’s instructions. Construction of libraries was prepared using the TruSeq Stranded Total RNA Sample Preparation Guide 15,031,048 Rev. E (Illumina, San Diego, USA), using 0.5–1 μg of total RNA input. The Novaseq 6000 platform was used and 100 bp paired-end reads were generated by Clinical Genomics at the University of Gothenburg (Gothenburg, Sweden).

Deshmukh M., Subhash S., Hu Z., Mohammad M., Jarneborn A., Pullerits R., Jin T, & Kopparapu P.K. (2023). Gene expression of S100a8/a9 predicts Staphylococcus aureus-induced septic arthritis in mice. Frontiers in Microbiology, 14, 1146694.

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Comprehensive Transcriptome and Small RNA Sequencing

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The 12 transcriptome libraries were prepared from 500 ng of total RNA using the TruSeq Stranded Total RNA Sample Preparation Kit with Ribo-Zero Gold (Illumina). Additionally, the 12 small RNA libraries were prepared from 1 μg from the same total RNA of transcriptome analysis, using the TruSeq Small RNA Library Prep Kit (Illumina). All the steps of the protocols were carried out without any modifications, with the exception of the cDNA construction purification step on the small RNA protocol, which was performed using 3% Agarose Gel Cassette for targets between 100 bp − 250 bp in a BluePippin system (Sage Science). The quality control of each library was performed using the 2100 Bioanalyzer System with the Agilent High Sensitivity DNA Kit (Agilent). The libraries were quantified via qPCR using a KAPA Library Quantification Kits for Illumina platforms (KAPA Biosystems). The Libraries were sequenced on a NextSeq. 500 sequencing system (Illumina), for the RNA-seq paired-end reads (2 × 75 bp) were obtained using NextSeq. 500/550 High Output v2 kit (150 cycles) and for the small RNA, single-end reads (1 × 50 bp) were obtained using a NextSeq. 500/550 High Output v2 kit (75 cycles).

Messias C.V., Loss-Morais G., Carvalho J.B., González M.N., Cunha D.P., Vasconcelos Z., Arge L.W., Farias-de-Oliveira D.A., Gerber A.L., Portari E.A., Ferreira N., Raphael L.M., Bonaldo M.C., Riederer I., Lopes Moreira M.E., Cotta-de-Almeida V., Vasconcelos A.T., Mendes-da-Cruz D.A, & Savino W. (2020). Zika virus targets the human thymic epithelium. Scientific Reports, 10, 1378.

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RNA-Seq Using Illumina Nextseq 500

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RNA-Seq was performed by GeneCore SU, University of Gothenburg, Sahlgrenska Academy, Gothenburg, Sweden (http://www.cgg.gu.se/genecore-su), using the Illumina Nextseq 500 platform (Illumina, San Diego, CA, USA). cDNA libraries were prepared from total RNA samples (≤1,500 ng), using the TruSeq Stranded Total RNA Sample Preparation Kit with Ribo-zero Gold (Illumina Inc.). Each library was paired-end sequenced (2 × 75 bp) using the Illumina Nextseq500 Kit High Output V2.

Mottram L., Lundgren A., Svennerholm A.M, & Leach S. (2021). A Systems Biology Approach Identifies B Cell Maturation Antigen (BCMA) as a Biomarker Reflecting Oral Vaccine Induced IgA Antibody Responses in Humans. Frontiers in Immunology, 12, 647873.

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Transcriptome Analysis of RNA-seq Data

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Preparation of libraries was carried out using the TruSeq Stranded Total RNA Sample Preparation Kit with Ribo-Zero Gold (Illumina, Inc., San Diego, CA), using 60–1000 ng of total RNA input. The Novaseq 6000 platform was used (Illumina, Inc., San Diego, CA) and 100 bp paired-end reads were generated by Clinical Genomics (Gothenburg, Sweden).
Adapters and low-quality tail were trimmed from reads prior to read alignment. Clean sequence reads aligned to the human genome were used to assemble transcripts, estimate the abundance of these transcripts and detect differential expression among samples. For mRNA and lncRNA analyses, the reference genome build GRCh38/hg38 was chosen as the annotation references. Fragments per kilo-base of exon per million fragments mapped (FPKM) of both lncRNAs and mRNAs in each sample were calculated based on the length of the fragments and reads count mapped to this fragment.

Torinsson Naluai Å., Östensson M., Fowler P.C., Abrahamsson S., Andersson B., Lassesson S., Jacobsson F., Oscarsson M., Bohman A., Harandi A.M, & Bende M. (2023). Transcriptomics unravels molecular changes associated with cilia and COVID-19 in chronic rhinosinusitis with nasal polyps. Scientific Reports, 13, 6592.

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Murine Hippocampus Transcriptome Profiling

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Total RNA was prepared from murine whole hippocampus using the Ambion mirVana miRNA Isolation Kit (AM1560; Invitrogen, Waltham, MA) and quantified using the Agilent 2100 Bioanalyzer nanochip (Agilent Technologies, Santa Clara, CA). Mean RNA integrity number for the samples was 9.2 (SD 0.5). Sixty libraries were prepared using the Illumina TruSeq Stranded Total RNA with Ribo-Zero gold sample preparation kit (Illumina, Inc., San Diego, CA), using 500 ng of input total RNA. Libraries were run on the Agilent 2100 Bioanalyzer High Sensitivity kit (Agilent Technologies) and combined in equimolar concentrations into three pools based on index color balance. Concentration of pools was measured with the KAPA Illumina Library Quantification Kit (KAPA Biosystems, Wilmington, MA). Pools were sequenced on Illumina HiSeq 2000 and 2500 sequencers with 100-bp Paired-End v3 SBS chemistry (Illumina) at the Iowa Institute of Human Genetics.

Michaelson J.J., Shin M.K., Koh J.Y., Brueggeman L., Zhang A., Katzman A., McDaniel L., Fang M., Pufall M, & Pieper A.A. (2017). Neuronal PAS Domain Proteins 1 and 3 Are Master Regulators of Neuropsychiatric Risk Genes. Biological psychiatry, 82(3), 213-223.

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RNA Sequencing Library Preparation

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RNA libraries were prepared at the Iowa Institute of Human Genetics (IIHG), Genomics Division, using the Illumina TruSeq Stranded Total RNA with Ribo-Zero gold sample preparation kit (Illumina, Inc., San Diego, CA). Library concentrations were measured using KAPA Illumina Library Quantification Kit (KAPA Biosystems, Wilmington, MA). Polled libraries were sequenced on Illumina NovaSeq6000 sequencer with 150-bp Paired-End chemistry (Illumina) at the IIHG core.

Vanrobeys Y., Mukherjee U., Langmack L., Bahl E., Lin L.C., Michaelson J.J., Abel T, & Chatterjee S. (2023). Mapping the spatial transcriptomic signature of the hippocampus during memory consolidation. bioRxiv.

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Transcriptome Sequencing Using Illumina NovaSeq

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RNA libraries were prepared at the Iowa Institute of Human Genetics (IIHG), Genomics Division, using the Illumina TruSeq Stranded Total RNA with Ribo-Zero gold sample preparation kit (Illumina, Inc., San Diego, CA). Library concentrations were measured using KAPA Illumina Library Quantification Kit (KAPA Biosystems, Wilmington, MA). Pooled libraries were sequenced on Illumina NovaSeq6000 sequencer with 150-bp Paired-End chemistry (Illumina) at the IIHG core.

Vanrobaeys Y., Mukherjee U., Langmack L., Beyer S.E., Bahl E., Lin L.C., Michaelson J.J., Abel T, & Chatterjee S. (2023). Mapping the spatial transcriptomic signature of the hippocampus during memory consolidation. Nature Communications, 14, 6100.

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RNA-seq Analysis of Mouse Models

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RNA quality was assessed and samples with RNA integrity number (RIN) > 8 were used to perform RNA-seq. RNA library preparation from WT mice (n = 8 samples, 4 males and 4 females) and 3g del/+ mice (n = 8 samples, 4 males and 4 females) for 3g del/+ mice and WT mice (n = 8 samples, 4 males and 4 females) and 16p11.2 del/+ mice (n = 8 samples, 4 males and 4 females) for 16p11.2 del/+ mice were prepared using the Illumina TruSeq Stranded Total RNA with Ribo-Zero gold sample preparation kit (Illumina, Inc., San Diego, CA). Pooled libraries were sequenced on Illumina NovaSeq 6000 sequencers with 100-bp Paired-End chemistry (Illumina). The dataset is available in the NCBI’s Gene Expression Omnibus repository, GEO Series accession GSE224750.

Abel T., Kim J., Vanrobaeys Y., Peterson Z., Kelvington B., Gaine M, & Nickl-Jockschat T. (2023). Dissecting 16p11.2 hemi-deletion to study sex-specific striatal phenotypes of neurodevelopmental disorders. Research Square.

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Transcriptome analysis of T. gondii

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Total RNA from T. gondii RH parasites treated with 24.2 nM trtE or 0.1% DMSO control was collected 4, 8, and 12 h post-treatment for RNA-Seq analysis. The details of the experimental analysis and results of the transcriptome are available in the GEO database (GSE140197). Briefly, the integrity and quantity of the total RNA samples were determined using a Fragment Analyzer™ (AATI, Ames, IA, USA). The RNA samples that satisfied the input requirement were used as input for the Illumina TruSeq Stranded Total RNA with Ribo-Zero® Gold sample preparation kit (Illumina, San Diego, CA, USA) following the manufacturer's instructions. The quality and molar concentration of the libraries were determined using the Fragment Analyzer. The libraries were pooled and sequenced using HiSeq 2500 High Output V4 chemistry (Illumina). The 75 million to 95 million reads per sample obtained were analyzed using the TopHat-Cufflinks pipeline. The expression level was estimated with Cuffdiff and represented as fragments per kilobase of exons per million mapped fragments (FPKM) to the T. gondii GT1 genome available on ToxoDB (Gajria et al., 2008 (link)) (http://toxodb.org). Genes were considered deferentially expressed when both the p-value and q-value, estimated based using a negative binomial model, were below a significance value of 0.05. The RNA-Seq experiment was conducted once.

Bowden G.D., Reis P.M., Rogers M.B., Bone Relat R.M., Brayton K.A., Wilson S.K., Di Genova B.M., Knoll L.J., Nepveux V F.J., Tai A.K., Ramadhar T.R., Clardy J, & O'Connor R.M. (2020). A conserved coccidian gene is involved in Toxoplasma sensitivity to the anti-apicomplexan compound, tartrolon E. International Journal for Parasitology: Drugs and Drug Resistance, 14, 1-7.

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RNA Sequencing of Hippocampal Transcripts

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RNA libraries from homecage and MWM mice were prepared at the Iowa Institute of Human Genetics (IIHG), Genomics Division, using the Illumina TruSeq Stranded Total RNA with Ribo-Zero gold sample preparation kit (Illumina, Inc., San Diego, CA). Library concentrations were measured with KAPA Illumina Library Quantification Kit (KAPA Biosystems, Wilmington, MA). Pooled libraries were sequenced on Illumina HiSeq 4000 sequencers with 150-bp Paired-End chemistry (Illumina) at the IIHG core.
The RNA seq data have been deposited in NCBI’s Gene Expression Omnibus and are accessible through GEO Series accession number GSE151681 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE151681). The code for analyses and figures related to RNA-seq data can be accessed through GitHub (github.com/ethanbahl/chatterjee2020_cbpkix).

Chatterjee S., Angelakos C.C., Bahl E., Hawk J.D., Gaine M.E., Poplawski S.G., Schneider-Anthony A., Yadav M., Porcari G.S., Cassel J.C., Giese K.P., Michaelson J.J., Lyons L.C., Boutillier A.L, & Abel T. (2020). The CBP KIX domain regulates long-term memory and circadian activity. BMC Biology, 18, 155.

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