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Nebnext mrna second strand synthesis module

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The NEBNext mRNA Second Strand Synthesis Module is a laboratory product that facilitates the synthesis of the second strand of complementary DNA (cDNA) from mRNA templates. It provides the necessary enzymes and reagents to convert single-stranded mRNA into double-stranded cDNA.

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Lab products found in correlation

Nextera xt dna library preparation kit, by Illumina (3 mentions) Trizol reagent, by Thermo Fisher Scientific (2 mentions) Ribo zero rrna removal kit, by Illumina (2 mentions) Hiseq 2500 system, by Illumina (1 mentions) Kapa library quantification kit, by Agilent Technologies (1 mentions) Bioanalyzer high sensitivity dna chip, by Agilent Technologies (1 mentions) Nebnext poly a mrna magnetic isolation module, by Illumina (1 mentions) Nextseq 500 instrument, by Illumina (1 mentions) Bioanalyzer, by Agilent Technologies (1 mentions)

4 protocols using nebnext mrna second strand synthesis module

1

RNA-seq of Adipocyte Transcriptome

Cited 1 time
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Total RNA from ~400 μl of thawed floated adipocytes was isolated in Trizol reagent (Invitrogen) following the manufacturer’s instructions. For RNA-seq library construction, mRNA was purified from 100 ng of total RNA using the Ribo-Zero rRNA removal kit (Epicentre) to deplete ribosomal RNA and converted into double stranded cDNA using NEBNext mRNA Second Strand Synthesis Module (E6111L). cDNA was subsequently tagmented and amplified for 12 cycles by using Nextera XT DNA Library Preparation Kit (Illumina FC-131). Sequencing libraries were analyzed Qubit and Agilent Bioanalyzer, pooled at a final loading concentration of 1.8pM, and sequenced on a NextSeq500. Sequencing reads were demultiplexed using bcl2fastq and aligned to the mm10 mouse genome using HISAT247 (link). PCR duplicates and low-quality reads were removed by Picard (https://broadinstitute.github.io/picard). Filtered reads were assigned to the annotated transcriptome and quantified using featureCounts48 (link).
Kazak L., Rahbani J.F., Samborska B., Lu G.Z., Jedrychowski M.P., Lajoie M., Zhang S., Ramsay L.C., Dou F.Y., Tenen D., Chouchani E.T., Dzeja P., Watson I.R., Tsai L., Rosen E.D, & Spiegelman B.M. (2019). Ablation of adipocyte creatine transport impairs thermogenesis and causes diet-induced obesity. Nature metabolism, 1(3), 360-370.
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2

RNA-seq of Adipocyte Transcriptome

Cited 1 time
Check if the same lab product or an alternative is used in the 5 most similar protocols
Total RNA from ~400 μl of thawed floated adipocytes was isolated in Trizol reagent (Invitrogen) following the manufacturer’s instructions. For RNA-seq library construction, mRNA was purified from 100 ng of total RNA using the Ribo-Zero rRNA removal kit (Epicentre) to deplete ribosomal RNA and converted into double stranded cDNA using NEBNext mRNA Second Strand Synthesis Module (E6111L). cDNA was subsequently tagmented and amplified for 12 cycles by using Nextera XT DNA Library Preparation Kit (Illumina FC-131). Sequencing libraries were analyzed Qubit and Agilent Bioanalyzer, pooled at a final loading concentration of 1.8pM, and sequenced on a NextSeq500. Sequencing reads were demultiplexed using bcl2fastq and aligned to the mm10 mouse genome using HISAT247 (link). PCR duplicates and low-quality reads were removed by Picard (https://broadinstitute.github.io/picard). Filtered reads were assigned to the annotated transcriptome and quantified using featureCounts48 (link).
Kazak L., Rahbani J.F., Samborska B., Lu G.Z., Jedrychowski M.P., Lajoie M., Zhang S., Ramsay L.C., Dou F.Y., Tenen D., Chouchani E.T., Dzeja P., Watson I.R., Tsai L., Rosen E.D, & Spiegelman B.M. (2019). Ablation of adipocyte creatine transport impairs thermogenesis and causes diet-induced obesity. Nature metabolism, 1(3), 360-370.
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3

RNA-Seq profiling of C. elegans HSF-1

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RNA was extracted by standard Trizol extraction techniques. Libraries were made using either NEBNext mRNA Second Strand Synthesis Module (E6111) followed by NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB-E7645) or NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (E7760) with the NEBNext Poly(A) mRNA Magnetic Isolation Module (NE7490) as per manufacturer’s protocols using half reactions. A total of 13 cycles of amplification was used for library enrichment; quality and size distribution of the libraries were ascertained by running on a Bioanalyzer High Sensitivity DNA Chip (Agilent (Santa Clara, USA), 5067–4626), and concentration was determined using KAPA Library Quantification Kit (KK4824). Libraries were sequenced on an Illumina HiSeq 2500 system by the Babraham Sequencing Facility. The experiment compared 3 independent replicates WT (N2) and hsf-1neuro (AGD1289) animals.
Chauve L., Hodge F., Murdoch S., Masoudzadeh F., Mann H.J., Lopez-Clavijo A.F., Okkenhaug H., West G., Sousa B.C., Segonds-Pichon A., Li C., Wingett S.W., Kienberger H., Kleigrewe K., de Bono M., Wakelam M.J, & Casanueva O. (2021). Neuronal HSF-1 coordinates the propagation of fat desaturation across tissues to enable adaptation to high temperatures in C. elegans. PLoS Biology, 19(11), e3001431.
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4

RNA-seq analysis of CoCl2-treated JAR cells

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Following 6 days of treatment with CoCl2 100 µM versus control media, total RNA was isolated from JAR cells as described above. Extracted RNA (300 ng) was treated with NEBNext rRNA Depletion Kit v2 (E7400X) and cDNA was generated using random hexamer priming and Maxima H Minus Reverse Transcriptase. cDNA was converted into double-stranded cDNA using NEBNext mRNA Second Strand Synthesis Module (E6111L) and sequencing libraries were generated by tagmentation using Nextera XT DNA Library Preparation Kit (Illumina FC-131) with 12 cycles of PCR amplification. Sequencing libraries were analyzed by Qubit and Agilent Bioanalyzer, pooled at a final concentration of 1.2 pM, and sequenced on an Illumina NextSeq500 instrument 36 × 8 × 36 read structure.
Ciampa E.J., Flahardy P., Srinivasan H., Jacobs C., Tsai L., Karumanchi S.A, & Parikh S.M. (2023). Hypoxia-inducible factor 1 signaling drives placental aging and can provoke preterm labor. eLife, 12, RP85597.
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