ultra directional mrnaseq kit  (New England Biolabs)


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    Name:
    NEBNext Ultra Directional RNA Library Prep Kit for Illumina
    Description:
    NEBNext Ultra Directional RNA Library Prep Kit for Illumina 96 rxns
    Catalog Number:
    e7420l
    Price:
    3801
    Size:
    96 rxns
    Category:
    mRNA Template Preparation for PCR
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    Structured Review

    New England Biolabs ultra directional mrnaseq kit
    NEBNext Ultra Directional RNA Library Prep Kit for Illumina
    NEBNext Ultra Directional RNA Library Prep Kit for Illumina 96 rxns
    https://www.bioz.com/result/ultra directional mrnaseq kit/product/New England Biolabs
    Average 90 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    ultra directional mrnaseq kit - by Bioz Stars, 2020-01
    90/100 stars

    Images

    1) Product Images from "Tri-methylation of histone H3 lysine 4 facilitates gene expression in ageing cells"

    Article Title: Tri-methylation of histone H3 lysine 4 facilitates gene expression in ageing cells

    Journal: eLife

    doi: 10.7554/eLife.34081

    Dynamics of H3K4me3 and expression in subsets of genes. ( A ) Genomic regions subject to age-linked CNV detected by comparison between input samples; these were excluded from ChIP analysis. ( B ) Scatter plot of log 2 -transformed mRNAseq read counts from protein coding genes defined in Figure 4A normalised for ORF length comparing log phase cells to 48-hr-aged cells. Data averaged across five replicates per condition. ( C ) Average profile of H3K4me3 ChIP signal for 1 kb either side of the TSS performed as in Figure 4C , for 300 genes least induced with age. ( D ) As ( C ) for 300 most induced genes. ( E ) Hierarchical clustering analysis of log 2 -transformed mRNA levels across age for the 4806 unambiguous genes defined in Figure 4A .
    Figure Legend Snippet: Dynamics of H3K4me3 and expression in subsets of genes. ( A ) Genomic regions subject to age-linked CNV detected by comparison between input samples; these were excluded from ChIP analysis. ( B ) Scatter plot of log 2 -transformed mRNAseq read counts from protein coding genes defined in Figure 4A normalised for ORF length comparing log phase cells to 48-hr-aged cells. Data averaged across five replicates per condition. ( C ) Average profile of H3K4me3 ChIP signal for 1 kb either side of the TSS performed as in Figure 4C , for 300 genes least induced with age. ( D ) As ( C ) for 300 most induced genes. ( E ) Hierarchical clustering analysis of log 2 -transformed mRNA levels across age for the 4806 unambiguous genes defined in Figure 4A .

    Techniques Used: Expressing, Chromatin Immunoprecipitation, Transformation Assay

    Related Articles

    Methylation Sequencing:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length). .. Libraries for ChIP‐seq and bisulfite sequencing were prepared as described (Feichtinger et al., ).

    Amplification:

    Article Title: Holo-Seq: single-cell sequencing of holo-transcriptome
    Article Snippet: The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). .. After Not I digestion, the library DNA was purified for USER digestion and PCR amplification (16 cycles) (more details in “Step-by-step Holo-Seq protocols” (Additional file )).

    Article Title: Transcriptome-wide Profiling of Cerebral Cavernous Malformations Patients Reveal Important Long noncoding RNA molecular signatures
    Article Snippet: RNA library preparation, quality control, and quantification 200 ng of total RNA per sample were utilized as input for rRNA depletion procedure with ‘NEBNext® rRNA Depletion Kit (Human/Mouse/Rat), 96 rxns’ (E6310X; New England Biolabs) followed by stranded cDNA library generation using ‘NEBNext® Ultra Directional RNA Library Prep Kit for Illumina’ (E7420L; New England Biolabs). .. All generated cDNA libraries were amplified by 11 cycles of final PCR.

    Construct:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA). .. The quality of the constructed cDNA library was determined using Agilent Bioanalyzer-High Sensitivity DNA Chip and Ribosome Integrity Numbers (RIN) were determined for all samples with a cutoff value of 10.

    Article Title: Holo-Seq: single-cell sequencing of holo-transcriptome
    Article Snippet: .. The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). .. After adaptor ligation, the cDNA fragments from the RNA carrier were removed by Not I digestion.

    Article Title: Defining the microbial transcriptional response to colitis through integrated host and microbiome profiling
    Article Snippet: .. Libraries were constructed using the NEBNext Ultra directional RNA library prep kit (New England Biolabs, Ipswich, MA, USA, E7420L). .. RNA sequencing was performed on the Illumina HiSeq 2000 (Illumina) resulting in an average of 45.38 M (17.75–84.46 M ) 100 bp paired-end reads per sample.

    Concentration Assay:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: RNA concentrations were measured in rRNA-depleted samples using NanoDrop 2000 UV-Vis Spectrophotometer with cutoff values for RNA concentration of 10 ng µl−1 . .. Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA).

    Article Title: ATXN1L, CIC, and ETS Transcription Factors Modulate Sensitivity to MAPK Pathway Inhibition
    Article Snippet: RNA sequencing libraries were prepared using a NEBNext Ultra Directional RNA Library Prep Kit for Illumina, NEB E7420. .. The concentration of each cDNA library was quantified with the KAPA Illumina ABI Quantification Kit (Kapa Biosystems).

    Modification:

    Article Title: Defining the microbial transcriptional response to colitis through integrated host and microbiome profiling
    Article Snippet: For simultaneous DNA and RNA extraction, pellets were dissolved in 500 μl extraction buffer (200 mm NaCl, 20 mm EDTA, 4 m guanidine thiocyanate and 1% (v/v) β-mercaptoethanol) and 210 μl 20% (w/v) SDS and lysed with a bead beater twice for 2 min using Lysing Matrix E tubes (MP Biomedicals, Santa Ana, CA, USA; procedure modified according to , ; ). .. Libraries were constructed using the NEBNext Ultra directional RNA library prep kit (New England Biolabs, Ipswich, MA, USA, E7420L).

    Ligation:

    Article Title: Holo-Seq: single-cell sequencing of holo-transcriptome
    Article Snippet: The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). .. After adaptor ligation, the cDNA fragments from the RNA carrier were removed by Not I digestion.

    Generated:

    Article Title: Transcriptome-wide Profiling of Cerebral Cavernous Malformations Patients Reveal Important Long noncoding RNA molecular signatures
    Article Snippet: RNA library preparation, quality control, and quantification 200 ng of total RNA per sample were utilized as input for rRNA depletion procedure with ‘NEBNext® rRNA Depletion Kit (Human/Mouse/Rat), 96 rxns’ (E6310X; New England Biolabs) followed by stranded cDNA library generation using ‘NEBNext® Ultra Directional RNA Library Prep Kit for Illumina’ (E7420L; New England Biolabs). .. All generated cDNA libraries were amplified by 11 cycles of final PCR.

    Sequencing:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA). .. Library sequencing was performed by the Nucleic Acids Research Facilities at Virginia Commonwealth University using Illumina HiSeq2000.

    Article Title: Holo-Seq: single-cell sequencing of holo-transcriptome
    Article Snippet: .. The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). .. After adaptor ligation, the cDNA fragments from the RNA carrier were removed by Not I digestion.

    Article Title: ATXN1L, CIC, and ETS Transcription Factors Modulate Sensitivity to MAPK Pathway Inhibition
    Article Snippet: RNA sequencing libraries were prepared using a NEBNext Ultra Directional RNA Library Prep Kit for Illumina, NEB E7420. .. Libraries were pooled for sequencing using the HiSeq 2500.

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Paragraph title: Sample preparation and sequencing ... Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    Article Title: Intrinsic cooperativity potentiates parallel cis-regulatory evolution
    Article Snippet: Libraries were prepared using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina E7420 (New England BioLabs). mRNA and library quality were assessed using a Bioanalyzer 2100 (Agilent). .. Sequencing was performed at the University of California, San Francisco Center for Advanced Technology on an Illumina HiSeq 4000.

    Article Title: Maternal Choline Supplementation during Normal Murine Pregnancy Alters the Placental Epigenome: Results of an Exploratory Study
    Article Snippet: Paragraph title: 2.4. Placental mRNA Sequencing and Data Analysis ... NEBNext Ultra Directional RNA Library Prep Kit (New England Biolabs, Ipswich, MA, USA, Catalog #: E7420) was used to make polyA+ RNAseq libraries, using 1 µg total RNA input.

    Article Title: Base J and H3.V Regulate Transcriptional Termination in Trypanosoma brucei
    Article Snippet: .. Stranded sequencing libraries were prepared using the NEBNext Ultra Directional Library Kit (E7420). .. Sequencing was per formed on an Illumina HiSeq 2000 sequencer using 50 bp reads.

    Article Title: Defining the microbial transcriptional response to colitis through integrated host and microbiome profiling
    Article Snippet: Paragraph title: Metagenomic and metatranscriptomic library preparation and sequencing ... Libraries were constructed using the NEBNext Ultra directional RNA library prep kit (New England Biolabs, Ipswich, MA, USA, E7420L).

    ChIP-sequencing:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Samples for ChIP‐seq were taken every 12 hr (1 × 107 viable cells for cell fixation and cell lysis; 1 × 106 cells for magnetic immunoprecipitation), for RNA‐seq every 24 hr (1 × 106 cells into 1 ml Trizol), and for whole‐genome bisulfite analysis at mid‐exponential and mid‐stationary phase (5 × 106 cells for DNA isolation; Supporting Information Figure 1). .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    DNA Extraction:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Samples for ChIP‐seq were taken every 12 hr (1 × 107 viable cells for cell fixation and cell lysis; 1 × 106 cells for magnetic immunoprecipitation), for RNA‐seq every 24 hr (1 × 106 cells into 1 ml Trizol), and for whole‐genome bisulfite analysis at mid‐exponential and mid‐stationary phase (5 × 106 cells for DNA isolation; Supporting Information Figure 1). .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    Nucleic Acid Electrophoresis:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA). .. The final cDNA products were purified with AMPure XP Beads (cat. no. A63880, Beckman Coulter, CA, USA) and band sizes were checked by gel electrophoresis.

    RNA Sequencing Assay:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: Paragraph title: Transcriptome analysis by RNA-seq ... Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA).

    Article Title: Heterochromatin-associated interactions of Drosophila HP1a with dADD1, HIPP1, and repetitive RNAs
    Article Snippet: .. An NEBNext ultradirectional RNA library kit (New England Biolabs, catalog no. E7420S) was used to make cDNA and RNA-seq libraries. .. Several minor modifications to the standard kit protocol were made based on (1) size (RNA fragmentation time was decreased from 15 to 5 min) and (2) the small amount of RNA (the number of PCR cycles were adjusted to 20 cycles in the USER excision and PCR library step).

    Article Title: ATXN1L, CIC, and ETS Transcription Factors Modulate Sensitivity to MAPK Pathway Inhibition
    Article Snippet: .. RNA sequencing libraries were prepared using a NEBNext Ultra Directional RNA Library Prep Kit for Illumina, NEB E7420. .. The concentration of each cDNA library was quantified with the KAPA Illumina ABI Quantification Kit (Kapa Biosystems).

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length). .. Libraries for ChIP‐seq and bisulfite sequencing were prepared as described (Feichtinger et al., ).

    Article Title: Maternal Choline Supplementation during Normal Murine Pregnancy Alters the Placental Epigenome: Results of an Exploratory Study
    Article Snippet: The following procedures were conducted by the Cornell RNA Sequencing Core (Ithaca, NY, USA). .. NEBNext Ultra Directional RNA Library Prep Kit (New England Biolabs, Ipswich, MA, USA, Catalog #: E7420) was used to make polyA+ RNAseq libraries, using 1 µg total RNA input.

    Article Title: BrAD-seq: Breath Adapter Directional sequencing: a streamlined, ultra-simple and fast library preparation protocol for strand specific mRNA library construction
    Article Snippet: .. Even allowing for the additional cost of consumables, chemical reagents and an extra 10% volume for reaction master mixes, this method provides a 20–40 fold cost reduction over available commercial strand-specific methods (e.g., NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® 96 reactions Cat. # E7420L, SureSelect Strand Specific RNA-Seq Library Preparation kit for 96 samples reactions Cat. # G9691A). .. Protocol development We had initially set out to modify a template switching protocol, but ended up making a discovery that would enable us to create arguably the cheapest and fastest RNA-seq protocol to date.

    Article Title: Base J and H3.V Regulate Transcriptional Termination in Trypanosoma brucei
    Article Snippet: Paragraph title: RNA-seq ... Stranded sequencing libraries were prepared using the NEBNext Ultra Directional Library Kit (E7420).

    Article Title: Defining the microbial transcriptional response to colitis through integrated host and microbiome profiling
    Article Snippet: Libraries were constructed using the NEBNext Ultra directional RNA library prep kit (New England Biolabs, Ipswich, MA, USA, E7420L). .. RNA sequencing was performed on the Illumina HiSeq 2000 (Illumina) resulting in an average of 45.38 M (17.75–84.46 M ) 100 bp paired-end reads per sample.

    Article Title: ESRP1 mutations cause hearing loss due to defects in alternative splicing that disrupt cochlear development
    Article Snippet: .. Total RNA (200ng) was used for poly A selected RNA-seq library preparation using the NEBNext® Ultra™ Directional RNA Library Prep Kit from Illumina® (mRNA) (New England Biolabs) (products: NEBNext® Poly(A) mRNA Magnetic Isolation Module (E7490S) and NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (E7420S). .. Biological replicates were individually barcoded, pooled, and sequenced on two lanes of a HiSeq 2000 platform for 100x2 bp paired-end RNA-seq at the Penn Next Generation Sequencing Core (NGSC) Facility.

    Isolation:

    Article Title: BrAD-seq: Breath Adapter Directional sequencing: a streamlined, ultra-simple and fast library preparation protocol for strand specific mRNA library construction
    Article Snippet: The cost of isolating mRNA and making strand-specific libraries with this method is extraordinarily low, with magnetic bead, dNTP, and enzyme costs totaling $2.96/sample including mRNA isolation or $1.98 if making libraries from mRNA. .. Even allowing for the additional cost of consumables, chemical reagents and an extra 10% volume for reaction master mixes, this method provides a 20–40 fold cost reduction over available commercial strand-specific methods (e.g., NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® 96 reactions Cat. # E7420L, SureSelect Strand Specific RNA-Seq Library Preparation kit for 96 samples reactions Cat. # G9691A).

    Article Title: ESRP1 mutations cause hearing loss due to defects in alternative splicing that disrupt cochlear development
    Article Snippet: .. Total RNA (200ng) was used for poly A selected RNA-seq library preparation using the NEBNext® Ultra™ Directional RNA Library Prep Kit from Illumina® (mRNA) (New England Biolabs) (products: NEBNext® Poly(A) mRNA Magnetic Isolation Module (E7490S) and NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (E7420S). .. Biological replicates were individually barcoded, pooled, and sequenced on two lanes of a HiSeq 2000 platform for 100x2 bp paired-end RNA-seq at the Penn Next Generation Sequencing Core (NGSC) Facility.

    Multiplex Assay:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: .. Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA). .. The final cDNA products were purified with AMPure XP Beads (cat. no. A63880, Beckman Coulter, CA, USA) and band sizes were checked by gel electrophoresis.

    Article Title: Transcriptome-wide Profiling of Cerebral Cavernous Malformations Patients Reveal Important Long noncoding RNA molecular signatures
    Article Snippet: RNA library preparation, quality control, and quantification 200 ng of total RNA per sample were utilized as input for rRNA depletion procedure with ‘NEBNext® rRNA Depletion Kit (Human/Mouse/Rat), 96 rxns’ (E6310X; New England Biolabs) followed by stranded cDNA library generation using ‘NEBNext® Ultra Directional RNA Library Prep Kit for Illumina’ (E7420L; New England Biolabs). .. Furthermore, one additional purification step was introduced at the end of the standard procedure, using 1x ‘Agencourt® AMPure® XP Beads’ (#A63881; Beckman Coulter, Inc.). cDNA libraries were barcoded by single indexing approach, using ‘NEBNext Multiplex Oligos for Illumina – Set 1’ (Index Primer 2, 3, 4, 5, 6, 7, and 12).

    Purification:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: For depletion of ribosomal RNA, all samples were treated with Illumina Ribo-zero Magnetic Kit for Bacteria (cat. no. MRZB12424, Roche, USA) and the rRNA-depleted samples were purified using Qiagen RNeasy MinElute Cleanup Kit (cat. no. 74204, Qiagen, CA, USA). .. Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA).

    Article Title: Holo-Seq: single-cell sequencing of holo-transcriptome
    Article Snippet: The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). .. After Not I digestion, the library DNA was purified for USER digestion and PCR amplification (16 cycles) (more details in “Step-by-step Holo-Seq protocols” (Additional file )).

    Article Title: Transcriptome-wide Profiling of Cerebral Cavernous Malformations Patients Reveal Important Long noncoding RNA molecular signatures
    Article Snippet: RNA library preparation, quality control, and quantification 200 ng of total RNA per sample were utilized as input for rRNA depletion procedure with ‘NEBNext® rRNA Depletion Kit (Human/Mouse/Rat), 96 rxns’ (E6310X; New England Biolabs) followed by stranded cDNA library generation using ‘NEBNext® Ultra Directional RNA Library Prep Kit for Illumina’ (E7420L; New England Biolabs). .. Furthermore, one additional purification step was introduced at the end of the standard procedure, using 1x ‘Agencourt® AMPure® XP Beads’ (#A63881; Beckman Coulter, Inc.). cDNA libraries were barcoded by single indexing approach, using ‘NEBNext Multiplex Oligos for Illumina – Set 1’ (Index Primer 2, 3, 4, 5, 6, 7, and 12).

    Polymerase Chain Reaction:

    Article Title: Holo-Seq: single-cell sequencing of holo-transcriptome
    Article Snippet: The sequencing library was constructed following the manufacturer’s protocol using a NEBNext kit (E7420). .. After Not I digestion, the library DNA was purified for USER digestion and PCR amplification (16 cycles) (more details in “Step-by-step Holo-Seq protocols” (Additional file )).

    Article Title: Heterochromatin-associated interactions of Drosophila HP1a with dADD1, HIPP1, and repetitive RNAs
    Article Snippet: An NEBNext ultradirectional RNA library kit (New England Biolabs, catalog no. E7420S) was used to make cDNA and RNA-seq libraries. .. Several minor modifications to the standard kit protocol were made based on (1) size (RNA fragmentation time was decreased from 15 to 5 min) and (2) the small amount of RNA (the number of PCR cycles were adjusted to 20 cycles in the USER excision and PCR library step).

    Article Title: Transcriptome-wide Profiling of Cerebral Cavernous Malformations Patients Reveal Important Long noncoding RNA molecular signatures
    Article Snippet: RNA library preparation, quality control, and quantification 200 ng of total RNA per sample were utilized as input for rRNA depletion procedure with ‘NEBNext® rRNA Depletion Kit (Human/Mouse/Rat), 96 rxns’ (E6310X; New England Biolabs) followed by stranded cDNA library generation using ‘NEBNext® Ultra Directional RNA Library Prep Kit for Illumina’ (E7420L; New England Biolabs). .. All generated cDNA libraries were amplified by 11 cycles of final PCR.

    cDNA Library Assay:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA). .. The quality of the constructed cDNA library was determined using Agilent Bioanalyzer-High Sensitivity DNA Chip and Ribosome Integrity Numbers (RIN) were determined for all samples with a cutoff value of 10.

    Article Title: ATXN1L, CIC, and ETS Transcription Factors Modulate Sensitivity to MAPK Pathway Inhibition
    Article Snippet: RNA sequencing libraries were prepared using a NEBNext Ultra Directional RNA Library Prep Kit for Illumina, NEB E7420. .. The concentration of each cDNA library was quantified with the KAPA Illumina ABI Quantification Kit (Kapa Biosystems).

    Article Title: Transcriptome-wide Profiling of Cerebral Cavernous Malformations Patients Reveal Important Long noncoding RNA molecular signatures
    Article Snippet: .. RNA library preparation, quality control, and quantification 200 ng of total RNA per sample were utilized as input for rRNA depletion procedure with ‘NEBNext® rRNA Depletion Kit (Human/Mouse/Rat), 96 rxns’ (E6310X; New England Biolabs) followed by stranded cDNA library generation using ‘NEBNext® Ultra Directional RNA Library Prep Kit for Illumina’ (E7420L; New England Biolabs). .. All steps were performed as recommended in user manualE7420 (Version 6.0_08-2015; NEB) except that all reactions were downscaled to 2/3 of initial volumes.

    Chromatin Immunoprecipitation:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA). .. The quality of the constructed cDNA library was determined using Agilent Bioanalyzer-High Sensitivity DNA Chip and Ribosome Integrity Numbers (RIN) were determined for all samples with a cutoff value of 10.

    RNA Extraction:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: For RNA extraction, the cells were centrifuged and lysed using TRI reagent (T9424–200 ml; Sigma‐Aldrich) following the manufacturer's instructions: phase separation was done by addition of chloroform and the aqueous phase collected. .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    Article Title: Defining the microbial transcriptional response to colitis through integrated host and microbiome profiling
    Article Snippet: For simultaneous DNA and RNA extraction, pellets were dissolved in 500 μl extraction buffer (200 mm NaCl, 20 mm EDTA, 4 m guanidine thiocyanate and 1% (v/v) β-mercaptoethanol) and 210 μl 20% (w/v) SDS and lysed with a bead beater twice for 2 min using Lysing Matrix E tubes (MP Biomedicals, Santa Ana, CA, USA; procedure modified according to , ; ). .. Libraries were constructed using the NEBNext Ultra directional RNA library prep kit (New England Biolabs, Ipswich, MA, USA, E7420L).

    Sample Prep:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Paragraph title: Sample preparation and sequencing ... Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    Next-Generation Sequencing:

    Article Title: ESRP1 mutations cause hearing loss due to defects in alternative splicing that disrupt cochlear development
    Article Snippet: Total RNA (200ng) was used for poly A selected RNA-seq library preparation using the NEBNext® Ultra™ Directional RNA Library Prep Kit from Illumina® (mRNA) (New England Biolabs) (products: NEBNext® Poly(A) mRNA Magnetic Isolation Module (E7490S) and NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina® (E7420S). .. Biological replicates were individually barcoded, pooled, and sequenced on two lanes of a HiSeq 2000 platform for 100x2 bp paired-end RNA-seq at the Penn Next Generation Sequencing Core (NGSC) Facility.

    Spectrophotometry:

    Article Title: Analysis of essential gene dynamics under antibiotic stress in Streptococcus sanguinis
    Article Snippet: RNA concentrations were measured in rRNA-depleted samples using NanoDrop 2000 UV-Vis Spectrophotometer with cutoff values for RNA concentration of 10 ng µl−1 . .. Actinomycin D (cat. no. A1410-2MG, Sigma-Aldrich, MO, USA) was used for RNA fragmentation and RNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit NEB (cat. no. E7420L, New England Biolabs, MA, USA) and NEBNext Multiplex Oligos for Illumina Index Primers Set 1 and set 2 (cat. nos E7335L and E7500L respectively, New England Biolabs, MA, USA).

    Immunoprecipitation:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Samples for ChIP‐seq were taken every 12 hr (1 × 107 viable cells for cell fixation and cell lysis; 1 × 106 cells for magnetic immunoprecipitation), for RNA‐seq every 24 hr (1 × 106 cells into 1 ml Trizol), and for whole‐genome bisulfite analysis at mid‐exponential and mid‐stationary phase (5 × 106 cells for DNA isolation; Supporting Information Figure 1). .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    Cell Counting:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: The total and viable cell count was analyzed twice daily with a ViCell (Beckmann Coulter). .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

    Lysis:

    Article Title: Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture, et al. Epigenetic regulation of gene expression in Chinese Hamster Ovary cells in response to the changing environment of a batch culture
    Article Snippet: Samples for ChIP‐seq were taken every 12 hr (1 × 107 viable cells for cell fixation and cell lysis; 1 × 106 cells for magnetic immunoprecipitation), for RNA‐seq every 24 hr (1 × 106 cells into 1 ml Trizol), and for whole‐genome bisulfite analysis at mid‐exponential and mid‐stationary phase (5 × 106 cells for DNA isolation; Supporting Information Figure 1). .. Libraries for RNA‐seq were prepared using NEBNext Ultra Directional RNA library prep Kits (E7420L), starting from total RNA according to the instructions and analyzed by Illumina HiSeq 2000 PE100 (pair‐end; 100 bp read length).

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    New England Biolabs ultra directional mrnaseq kit
    Dynamics of H3K4me3 and expression in subsets of genes. ( A ) Genomic regions subject to age-linked CNV detected by comparison between input samples; these were excluded from ChIP analysis. ( B ) Scatter plot of log 2 -transformed <t>mRNAseq</t> read counts from protein coding genes defined in Figure 4A normalised for ORF length comparing log phase cells to 48-hr-aged cells. Data averaged across five replicates per condition. ( C ) Average profile of H3K4me3 ChIP signal for 1 kb either side of the TSS performed as in Figure 4C , for 300 genes least induced with age. ( D ) As ( C ) for 300 most induced genes. ( E ) Hierarchical clustering analysis of log 2 -transformed mRNA levels across age for the 4806 unambiguous genes defined in Figure 4A .
    Ultra Directional Mrnaseq Kit, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 90/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Dynamics of H3K4me3 and expression in subsets of genes. ( A ) Genomic regions subject to age-linked CNV detected by comparison between input samples; these were excluded from ChIP analysis. ( B ) Scatter plot of log 2 -transformed mRNAseq read counts from protein coding genes defined in Figure 4A normalised for ORF length comparing log phase cells to 48-hr-aged cells. Data averaged across five replicates per condition. ( C ) Average profile of H3K4me3 ChIP signal for 1 kb either side of the TSS performed as in Figure 4C , for 300 genes least induced with age. ( D ) As ( C ) for 300 most induced genes. ( E ) Hierarchical clustering analysis of log 2 -transformed mRNA levels across age for the 4806 unambiguous genes defined in Figure 4A .

    Journal: eLife

    Article Title: Tri-methylation of histone H3 lysine 4 facilitates gene expression in ageing cells

    doi: 10.7554/eLife.34081

    Figure Lengend Snippet: Dynamics of H3K4me3 and expression in subsets of genes. ( A ) Genomic regions subject to age-linked CNV detected by comparison between input samples; these were excluded from ChIP analysis. ( B ) Scatter plot of log 2 -transformed mRNAseq read counts from protein coding genes defined in Figure 4A normalised for ORF length comparing log phase cells to 48-hr-aged cells. Data averaged across five replicates per condition. ( C ) Average profile of H3K4me3 ChIP signal for 1 kb either side of the TSS performed as in Figure 4C , for 300 genes least induced with age. ( D ) As ( C ) for 300 most induced genes. ( E ) Hierarchical clustering analysis of log 2 -transformed mRNA levels across age for the 4806 unambiguous genes defined in Figure 4A .

    Article Snippet: 500 ng RNA was used to prepare libraries using the NEBNext Ultra Directional mRNAseq kit (NEB E7420, E7490, E7335, E7500, E7710) as described with modifications: Reagent volumes for RT, second strand synthesis, tailing and ligation were reduced by 50%, while libraries were amplified for 12 cycles using 2 µl each primer per reaction before two rounds of AMPure bead purification as described in the kit at 0.9x ratio prior to quality control using a Bioanalyzer and quantification usinng KAPA Biosystems kit KK4824.

    Techniques: Expressing, Chromatin Immunoprecipitation, Transformation Assay