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Nebnext small rna kit

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The NEBNext Small RNA kit is a laboratory product designed for the preparation of small RNA libraries for next-generation sequencing. It provides a streamlined workflow for the isolation, purification, and adapter ligation of small RNA molecules from various sample types.

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Lab products found in correlation

Agilent 2100 bioanalyzer, by Agilent Technologies (3 mentions) Hiseq 4000, by Illumina (2 mentions) Hiseq 2500 platform, by Illumina (2 mentions) Mirneasy serum plasma kit, by Qiagen (2 mentions) Hiseq 1000, by Illumina (2 mentions) Hiseq 2500, by Illumina (2 mentions) Trizol reagent, by Thermo Fisher Scientific (2 mentions) Allprep ffpe kit, by Qiagen (1 mentions) Pippin prep, by Sage Science (1 mentions) Agilent bioanalyzer, by Agilent Technologies (1 mentions) Rneasy kit, by Qiagen (1 mentions) Novaseq, by Illumina (1 mentions) Smarter universal low input rna kit, by Takara Bio (1 mentions) Nextseq, by Illumina (1 mentions) Nextseq platform, by Illumina (1 mentions) Rnase free dnase, by Qiagen (1 mentions) Qiaseq mirna library qc spike ins, by Qiagen (1 mentions) Truseq small rna library prep kit, by New England Biolabs (1 mentions) Nextseq 500, by Illumina (1 mentions) Mirvana mirna isolation kit, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

NEBNext Multiplex Small RNA Library Prep kit (42 citations) NEBNext Small RNA Library Prep Kit (29 citations) NEBNext Small RNA library preparation kit (25 citations) NEBNext Small RNA Library kit (12 citations) NEBNext Multiplex Small RNA Kit (7 citations) NEBNext kits (3 citations) NEBNext Multiplex Small RNA Library Preparation Kit (3 citations) NEBNext Multiplex Small RNA Library kit (2 citations) NEBNext Small RNA Sequencing Library Preparation kit (2 citations)

14 protocols using nebnext small rna kit

1

BoRed-Seq and m7G Immunoprecipitation Protocol

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The detailed protocol of all the procedures required to perform BoRed-Seq and m7G RNA immunoprecipitation experiments on small RNAs is described in Methods S1. Single-end 50-bp stranded smallRNA libraries were prepared using the NEBNext SmallRNA kit (NEB) according to the manufacturer’s recommendations and sequenced on a HiSeq 4000 (Illumina).
Pandolfini L., Barbieri I., Bannister A.J., Hendrick A., Andrews B., Webster N., Murat P., Mach P., Brandi R., Robson S.C., Migliori V., Alendar A., d’Onofrio M., Balasubramanian S, & Kouzarides T. (2019). METTL1 Promotes let-7 MicroRNA Processing via m7G Methylation. Molecular Cell, 74(6), 1278-1290.e9.
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2

Gallbladder Tissue RNA-seq Protocol

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Formalin-fixed, paraffin-embedded (FFPE) gallbladder tissue specimens were obtained from 98 patients in total (n = 31 GS; n = 35 Dys; n = 32 GBC). RNA was extracted from FFPE sections using the AllPrep FFPE kit following Qiagen’s recommendations, and RNA quality was controlled (High Sensitivity Genomic DNA, Advanced Analytical, United States, and FFPE quality control kits, Illumina).
The NEBNext Small RNA kit (NEB) was used to produce RNA sequencing libraries, which were sequenced on the HiSeq 2500 platform (Illumina, San Diego, CA, USA) to an average depth of 18 M reads per sample. The applied RNA sequencing protocol has been previously described in detail [18 (link)]. Briefly, our protocol enabled us to capture lncRNA mapped fragments in the size range up to 47 base pairs. First, reads from the HiSeq 2500 platform were adapter-trimmed (AdapterRemoval v2.1.7) [19 (link)]. Then, adapter-trimmed reads were mapped to the human genome (hg38) by a Bowtie2 v2.2.9 aligner [20 (link)]. HTSeq was used to count reads mapped to lncRNA regions in GENCODE v26 annotations [21 (link),22 (link)].
Blandino A., Scherer D., Rounge T.B., Umu S.U., Boekstegers F., Barahona Ponce C., Marcelain K., Gárate-Calderón V., Waldenberger M., Morales E., Rojas A., Munoz C., Retamales J., de Toro G., Barajas O., Rivera M.T., Cortés A., Loader D., Saavedra J., Gutiérrez L., Ortega A., Bertrán M.E., Gabler F., Campos M., Alvarado J., Moisán F., Spencer L., Nervi B., Carvajal-Hausdorf D.E., Losada H., Almau M., Fernández P., Gallegos I., Olloquequi J., Fuentes-Guajardo M., Gonzalez-Jose R., Bortolini M.C., Gallo C., Linares A.R., Rothhammer F, & Lorenzo Bermejo J. (2022). Identification of Circulating lncRNAs Associated with Gallbladder Cancer Risk by Tissue-Based Preselection, Cis-eQTL Validation, and Analysis of Association with Genotype-Based Expression. Cancers, 14(3), 634.
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3

miRNA Extraction and Sequencing from Serum

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We extracted RNA from 400 µL serum using phenol-chloroform and miRNeasy Serum/Plasma kit (Qiagen, Valencia, CA). We performed size selection using a 3% Agarose Gel Cassette (Cat. No CSD3010) on a Pippin Prep (Sage Science) with a cut size optimized to cover RNA molecules from 17 to 47 nt in length. Libraries were prepared with the NEBNext Small RNA kit (NEB, Ipswich, MA) and sequenced on a HiSeq 2500 platform to on average 18 million sequences per sample (Illumina, San Diego, CA).
Umu S.U., Langseth H., Zuber V., Helland Å., Lyle R, & Rounge T.B. (2022). Serum RNAs can predict lung cancer up to 10 years prior to diagnosis. eLife, 11, e71035.
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4

Ribosome 2'-O-methylation profiling by RiboMethSeq

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RiboMethSeq analysis of rRNA 2’-O-methylation was performed as described in [60 (link)]. Briefly, ~100 ng of total RNA extracted from biological material was fragmented in bicarbonate buffer at pH 9.3 at 96°C, fragmentation time was adapted to obtain short RNA fragments of ~20–30 nt in length. Fragmented RNA was end-repaired, to insure compatibility with adapter ligation. Library preparation was done using NEBNext Small RNA kit, according to manufacturer’s recommendations. Barcoded libraries were loaded to the flow-cell and sequenced on Illumina HiSeq 1000. Raw reads were trimmed and aligned to the reference. Bioinformatics analysis was performed as described [60 (link)]. The complete data set is available at the European Nucleotide Archive under accession n°PRJEB42253.
Delhermite J., Tafforeau L., Sharma S., Marchand V., Wacheul L., Lattuca R., Desiderio S., Motorin Y., Bellefroid E, & Lafontaine D.L. (2022). Systematic mapping of rRNA 2’-O methylation during frog development and involvement of the methyltransferase Fibrillarin in eye and craniofacial development in Xenopus laevis. PLoS Genetics, 18(1), e1010012.
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5

Illumina-based small RNA sequencing

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Libraries for CAInput, FAInput, CAIP, and FAIP were prepared using the NEBNext Small RNA kit (NEB, Ipswich, MA) by the Genomic Sequencing and Analysis Facility (GSAF) at the University of Texas at Austin. Sequencing was performed on an Illumina HiSeq 4000 to yield 75-bp reads with an average read depth of 31 M reads for pull-downs and 56 M reads for total RNA samples. Raw sequencing results are available in the NCBI GEO database (Accession number: GSE148377).
Gonzalez-Rivera J.C., Sherman M.W., Wang D.S., Chuvalo-Abraham J.C., Hildebrandt Ruiz L, & Contreras L.M. (2020). RNA oxidation in chromatin modification and DNA-damage response following exposure to formaldehyde. Scientific Reports, 10, 16545.
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6

High-throughput sequencing of coding and non-coding RNA

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The coding RNA libraries for high-throughput sequencing were generated using NEBNext RNA Ultra II reagents (Ipswich, Massachusetts) and the generated libraries were paired-end sequenced on the NovaSeq platform at 150 cycles. ncRNA libraries were prepared from total RNA using NEBNext smallRNA kit (New England BioLabs, Ipswich, MA). The libraries quality control metrics were determined using agilent bioanalyzer and the generated libraries were sequenced on the NovaSeq platform. The integrity of the generated libraries for both the coding or noncoding libraries were determined using Agilent 2100 bioanalyzer.
Ramamoorthi Elangovan V., Saadat N., Ghnenis A., Padmanabhan V, & Vyas A.K. (2023). Developmental programming: adverse sexually dimorphic transcriptional programming of gestational testosterone excess in cardiac left ventricle of fetal sheep. Scientific Reports, 13, 2682.
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7

Ribosome Profiling and mRNA-Seq Workflow

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Ribosome profiling was performed by digesting subcellular fractions with micrococcal nuclease, pelleting ribosomes, and purifying the ribosome-protected RNA footprint by gel electrophoresis. Deep sequencing libraries were prepared using the NEBNext Small RNA kit (New England Biolabs). For mRNA-seq, mRNAs were enriched by rRNA depletion, then fragmented for library preparation. All sequencing was performed on the Illumina HiSeq 2500. Experimental duplicates were performed for each time point.
Reid D.W., Chen Q., Tay A.S., Shenolikar S, & Nicchitta C.V. (2014). The Unfolded Protein Response Triggers Selective mRNA Release From the Endoplasmic Reticulum. Cell, 158(6), 1362-1374.
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8

RNA Isolation, Sequencing, and Analysis Protocol

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Tissues were homogenized in Trizol reagent (Life Technologies, Carlsbad, CA) and total RNA was isolated following the manufacturer’s recommendations. RNA was purified using RNeasy kit (Qiagen, Germantown, MD) with contaminating DNA removed using RNAse free DNAse. The purity and RNA integrity, was evaluated using Agilent 2100 bioanalyzer (Agilent Technologies, Santa Clara, CA). For RNA sequencing, total RNA libraries were prepared with SMARTer universal low input RNA kit (Takara, Mountain View, CA) following ribosome depletion. Paired-end sequencing was performed on a NovaSeq (Illumina, San Diego, CA) for 150 cycles (75 base pairs on each paired end). Libraries for ncRNA (small RNA) were prepared from total RNA and sequencing libraries generated using the NEBNext smallRNA kit (New England Biolabs, Ipswich, MA). Sequencing of the libraries was accomplished on a Nextseq (Illumina, San Diego, CA) instrument. Extraction of small RNA from total RNA, sample quality evaluation, library preparation, and post-library quality control metrics were performed at the University of Michigan Advanced Genomics Core.
Puttabyatappa M., Saadat N., Elangovan V.R., Dou J., Bakulski K, & Padmanabhan V. (2022). Developmental Programming: Impact of Prenatal Bisphenol-A Exposure on Liver and Muscle Transcriptome of Female Sheep. Toxicology and applied pharmacology, 451, 116161.
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9

Profiling Serum Small RNAs by Sequencing

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RNA was extracted from 400 µL serum using phenol–chloroform and miRNeasy Serum/Plasma kit (Qiagen, Valencia, CA, USA). Libraries were prepared with the NEBNext Small RNA kit (NEB, Ipswich, MA, USA) and sequenced on a HiSeq 2500 (Illumina, San Diego, CA, USA) as previously described (Umu et al., 2018). RNA profiles from the JSB healthy donor samples show high RNA diversity, including sncRNAs and also fragments of longer RNAs (lncRNAs and mRNAs) (Rounge et al., 2015; Umu et al., 2018).
Umu S.U., Langseth H., Keller A., Meese E., Helland Å., Lyle R, & Rounge T.B. (2020). A 10‐year prediagnostic follow‐up study shows that serum RNA signals are highly dynamic in lung carcinogenesis. Molecular Oncology, 14(2), 235-247.
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10

Small RNA Extraction and Sequencing

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Adipose tissue was homogenized with liquid nitrogen and dissolved in Trizol reagent (Life Technologies, Carlsbad, CA) and total RNA isolated as per manufacturer’s recommendations. Residual DNA was removed using the RNeasy binding column, treating with RNAse free DNAse (Qiagen, Germantown, MD) then eluting RNA in nuclease free water. RNA purity and integrity were measured with the Agilent 2100 bioanalyzer (Agilent Technologies, Santa Clara, CA). Libraries for ncRNA were prepared using the NEBNext smallRNA kit (New England BioLabs, Ipswich, MA) at the University of Michigan Advanced Genomics Core as per manufacturer’s recommendations. Sequencing was performed on an Illumina Nextseq platform.
Dou J., Thangaraj S.V., Puttabyatappa M., Elangovan V.R., Bakulski K, & Padmanabhan V. (2023). Developmental Programming: adipose depot-specific regulation of non-coding RNAs and their relation to coding RNA expression in prenatal testosterone and prenatal bisphenol-A -treated female sheep. Molecular and cellular endocrinology, 564, 111868.
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