multiplex oligos oligonucleotides  (New England Biolabs)


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    Name:
    NEBNext Multiplex Oligos for Illumina Index Primers Set 1
    Description:
    NEBNext Multiplex Oligos for Illumina Index Primers Set 1 96 rxns
    Catalog Number:
    e7335l
    Price:
    385
    Size:
    96 rxns
    Category:
    Probes and Primers
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    Structured Review

    New England Biolabs multiplex oligos oligonucleotides
    NEBNext Multiplex Oligos for Illumina Index Primers Set 1
    NEBNext Multiplex Oligos for Illumina Index Primers Set 1 96 rxns
    https://www.bioz.com/result/multiplex oligos oligonucleotides/product/New England Biolabs
    Average 99 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    multiplex oligos oligonucleotides - by Bioz Stars, 2020-08
    99/100 stars

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    Related Articles

    Multiplex Assay:

    Article Title: UNC-16 (JIP3) Acts Through Synapse-Assembly Proteins to Inhibit the Active Transport of Cell Soma Organelles to Caenorhabditis elegans Motor Neuron Axons
    Article Snippet: .. To produce libraries for whole-genome sequencing, we quantified the DNA concentrations using the Qubit fluorimeter (Invitrogen) and the Qubit dsDNA BR Assay Kit (Invitrogen), sheared the DNAs to ∼350 bp by transferring 1080 ng of each DNA in 60 μl of water to Covaris Microtubes using two 25-sec cycles in a Covaris S2 at 4°, and then followed the manufacturer’s instructions for the NEBNext Ultra DNA Library Prep Kit for Illumina (NEB E7370S) and NEBNext Multiplex Oligos for Illumina (Index Primers set 1 and set 2; E7335S and E7500S). .. We analyzed the average size and concentration of the fragments in each library using the Agilent 2200 Tape Station and adjusted the concentration of each library to equal that of the lowest concentration library.

    Article Title: Histone H3 lysine 4 methylation signature associated with human undernutrition
    Article Snippet: .. ERCC spike-in RNA (Illumina) was added to aliquots of total RNA per instructions of the manufacturer, ribosomal RNA was then depleted using the RiboZero gold kit and libraries were constructed using the NEBNext Ultradirectional RNA Lib Prep Kit (Cat #E74205) and NEBNext Multiplex Oligos (Cat#E73355). ..

    Article Title: Hamp1 mRNA and plasma hepcidin levels are influenced by sex and strain but do not predict tissue iron levels in inbred mice
    Article Snippet: .. Libraries were prepared by using the NEBNext Poly(A) mRNA Magnetic Isolation Module, the NEBNext Ultra RNA Library Prep Kit for Illumina, and the NEBNext Multiplex Oligos for Illumina (Ipswich, MA) according to the manufacturer’s instructions. .. In brief, 500 ng of liver total RNA were mixed with prewashed NEBNext Oligo d(T)25 beads, placed on the magnetic rack, and eluted with 0.1× Invitrogen Tris-EDTA buffer (Carlsbad, CA) to isolate poly-A mRNA.

    Article Title: Quantitative Shearing Linear Amplification Polymerase Chain Reaction: An Improved Method for Quantifying Lentiviral Vector Insertion Sites in Transplanted Hematopoietic Cell Systems
    Article Snippet: .. The other primer for the nested PCR binds to the adaptor (13 bp complementary) and is provided in the NEBNext Multiplex Oligos for Illumina kit. .. The final PCR product was size-selected for the 250–800 bp fragments on a 2% E-gel (Life Technologies), was purified, and sequenced on a MiSeq instrument (Illumina).

    Article Title: Utilization of Tissue Ploidy Level Variation in de Novo Transcriptome Assembly of Pinus sylvestris
    Article Snippet: .. Libraries were indexed using NEBNext Multiplex Oligos for Illumina, Single Index Set 1. .. RNA library concentrations were quantified using NEBNext Library Quant Kit for Illumina and LightCycler 480 (Roche).

    Article Title: Tumor Cells are Dislodged into the Pulmonary Vein During Lobectomy
    Article Snippet: .. Library preparation was done with the NEBNext DNA Library Prep Master Mix Set for Illumina (NEB) and barcoded with the NEBNext Multiplex Oligos for Illumina (NEB). .. Sequencing of barcoded pools was performed with paired-end 150 reads using the Illumina MiSeq and data were analyzed using a combination of the Cloud-Based DNAseq Sequence Variant Analysis (Qiagen).

    Article Title: Dysregulation of the epigenetic landscape of normal aging in Alzheimer’s disease
    Article Snippet: .. Libraries were multiplexed using NEBNext Multiplex Oligos for Illumina (dual index primers) and single-ended sequenced (75 bp) on the NextSeq 500 platform (Illumina) in accordance with the manufacturer’s protocol. .. ChIP-seq tags generated with the NextSeq 500 platform were de-multiplexed with the bcl2fastq utility and aligned to the human reference genome (assembly NCBI37/hg19) using Bowtie v1.1.1 , allowing up to two mismatches per sequencing tag (parameters: −m 1–best).

    Article Title: Mapping Cellular Reprogramming via Pooled Overexpression Screens with Paired Fitness and Single Cell RNA-Sequencing Readout
    Article Snippet: .. Next Multiplex Oligos for Illumina (New England Biolabs) Index primers were used to attach Illumina adapters and indices to the samples. .. Next Multiplex Oligos for Illumina (New England Biolabs) Index primers were used to attach Illumina adapters and indices to the samples.

    Isolation:

    Article Title: Hamp1 mRNA and plasma hepcidin levels are influenced by sex and strain but do not predict tissue iron levels in inbred mice
    Article Snippet: .. Libraries were prepared by using the NEBNext Poly(A) mRNA Magnetic Isolation Module, the NEBNext Ultra RNA Library Prep Kit for Illumina, and the NEBNext Multiplex Oligos for Illumina (Ipswich, MA) according to the manufacturer’s instructions. .. In brief, 500 ng of liver total RNA were mixed with prewashed NEBNext Oligo d(T)25 beads, placed on the magnetic rack, and eluted with 0.1× Invitrogen Tris-EDTA buffer (Carlsbad, CA) to isolate poly-A mRNA.

    Transferring:

    Article Title: UNC-16 (JIP3) Acts Through Synapse-Assembly Proteins to Inhibit the Active Transport of Cell Soma Organelles to Caenorhabditis elegans Motor Neuron Axons
    Article Snippet: .. To produce libraries for whole-genome sequencing, we quantified the DNA concentrations using the Qubit fluorimeter (Invitrogen) and the Qubit dsDNA BR Assay Kit (Invitrogen), sheared the DNAs to ∼350 bp by transferring 1080 ng of each DNA in 60 μl of water to Covaris Microtubes using two 25-sec cycles in a Covaris S2 at 4°, and then followed the manufacturer’s instructions for the NEBNext Ultra DNA Library Prep Kit for Illumina (NEB E7370S) and NEBNext Multiplex Oligos for Illumina (Index Primers set 1 and set 2; E7335S and E7500S). .. We analyzed the average size and concentration of the fragments in each library using the Agilent 2200 Tape Station and adjusted the concentration of each library to equal that of the lowest concentration library.

    Construct:

    Article Title: Histone H3 lysine 4 methylation signature associated with human undernutrition
    Article Snippet: .. ERCC spike-in RNA (Illumina) was added to aliquots of total RNA per instructions of the manufacturer, ribosomal RNA was then depleted using the RiboZero gold kit and libraries were constructed using the NEBNext Ultradirectional RNA Lib Prep Kit (Cat #E74205) and NEBNext Multiplex Oligos (Cat#E73355). ..

    Sequencing:

    Article Title: UNC-16 (JIP3) Acts Through Synapse-Assembly Proteins to Inhibit the Active Transport of Cell Soma Organelles to Caenorhabditis elegans Motor Neuron Axons
    Article Snippet: .. To produce libraries for whole-genome sequencing, we quantified the DNA concentrations using the Qubit fluorimeter (Invitrogen) and the Qubit dsDNA BR Assay Kit (Invitrogen), sheared the DNAs to ∼350 bp by transferring 1080 ng of each DNA in 60 μl of water to Covaris Microtubes using two 25-sec cycles in a Covaris S2 at 4°, and then followed the manufacturer’s instructions for the NEBNext Ultra DNA Library Prep Kit for Illumina (NEB E7370S) and NEBNext Multiplex Oligos for Illumina (Index Primers set 1 and set 2; E7335S and E7500S). .. We analyzed the average size and concentration of the fragments in each library using the Agilent 2200 Tape Station and adjusted the concentration of each library to equal that of the lowest concentration library.

    Nested PCR:

    Article Title: Quantitative Shearing Linear Amplification Polymerase Chain Reaction: An Improved Method for Quantifying Lentiviral Vector Insertion Sites in Transplanted Hematopoietic Cell Systems
    Article Snippet: .. The other primer for the nested PCR binds to the adaptor (13 bp complementary) and is provided in the NEBNext Multiplex Oligos for Illumina kit. .. The final PCR product was size-selected for the 250–800 bp fragments on a 2% E-gel (Life Technologies), was purified, and sequenced on a MiSeq instrument (Illumina).

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    New England Biolabs universal pcr primers
    Mapping nucleosome distributions following KSHV reactivation using mTSS-seq technique and mTSS-seq validation A. Experimental design for mapping nucleosome distributions following KSHV reactivation using our newly developed mTSS-seq technique. B. Validation of mTSS-seq using quantitative <t>PCR</t> for both on-target regions of the genome (within the sequence-capture region, two-kb region centered on a TSS) and off-target regions of the genome (not within the sequence-capture region, outside the two-region centered on a TSS). Quantitative PCR was performed for on- and off-target regions of the genome for both RHOC and ITGA4. The y-axis shows C t values. On average, C t values between the on- and off-targets of the sequence-captured libraries differ by 10.5 cycles. C. The periodic occurrence of AA/TT/AT/TA dinucleotides was calculated for all nucleosomal-sized fragments (147-148 bp) at the 0 hour time point. The x-axis represents the distance from the dyad axis. The y-axis is the frequency of AA/TT/AT/TA dinucleotides. An A/T-containing dinucleotide periodicity is seen every 10 bp at the 0 hour time point. D. Alignment of the midpoint fragments (purple) from mTSS-seq to the human genome shown in the UCSC Genome Browser ( http://genome.ucsc.edu ). Zooming in 5000X on human chromosome 2 to a six-kb window with two-kb of midpoint fragments at 0 hour time point (purple lines), along with the sequence-capture <t>oligos</t> and previously-published human-nucleosome distribution map for cell line GM18508 (red lines, (Gaffney et al., 2012)) at the TSSs of PUS10 and PEX13. The x-axis is genomic location. The y-axis is scaled reads per million.
    Universal Pcr Primers, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs nebnext multiplex oligos for illumina
    The newly developed mTSS-Capture method combined with paired-end sequencing maps genome-wide nucleosome distribution in primary patient samples and identifies bona fide nucleosome characteristics, concordant with other human nucleosome mapping studies A . Work-flow of the mTSS-seq method. Following MNase digestion using a titration of MNase, populations of mononucleosomally protected DNA and subnucleosomal fragments are isolated, and prepared as libraries for <t>Illumina</t> sequencing. Solution-based sequence capture is performed using biotinylated <t>oligos,</t> enabling the enrichment of fragments within 2kb of each transcription start site in the human genome. Paired-end 50bp sequencing was then performed on each index. B . Alignment of the mTSS-seq midpoints to the human genome using the UCSC genome browser for LAC patient #4137 Normal tissue is shown for chr11, hg19 ( http://genome.ucsc.edu ). Zooming in twice at 100X allows for further visualization of the sequence capture oligos surrounding the TSS in a 500kb and a 5kb region showing the ATM locus. C . Averaged, normalized reads per million (y-axis) from mTSS-seq plotted as fragments (gray) and midpoints (black), centered on and surrounding 2kb of the TSS for ~22,000 open reading frames in hg19 (x-axis). DNase I-hypersensitivity (GSM736580; green) and RNA polymerase II from ChIP-seq (GSM935299; blue) data from A549 cells are shown. (D) LAC patient 4137 Normal nucleosomal midpoints (blue track) were plotted in the UCSC genome browser against the published human lymphocyte nucleosome distribution maps by Gaffney et. al. (green track) for the ZNF451 and CCDC97 loci. Sequence capture oligos and corresponding RefSeq gene models are shown for each locus. Correlations are shown for ZNF451 and CCDC87, respectively.
    Nebnext Multiplex Oligos For Illumina, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 309 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/nebnext multiplex oligos for illumina/product/New England Biolabs
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    New England Biolabs nebnext index primer for illumina
    Schistosomula excrete/secrete extracellular miRNAs. Supernatant from 72 h in vitro cultured schistosomula was separated into EV-enriched and EV-depleted fractions (n=3) by preparatory ultracentrifugation. Total RNA from each fraction was extracted using the miRNeasy kit (Qiagen) and prepared for RNA-seq using the <t>NEBNext</t> Small RNA (for <t>Illumina</t> HiSeq sequencing) kit. To identify sma-miRNAs in our samples, the miRDeep2 package was utilized (29). Only miRNAs with at least 10 reads in 2 out of the 3 biological replicates (in at least one of the fractions, EV-enriched OR EV-depleted) were considered in the study. A total of 205 putative sma-miRNAs passed this criterion. sma-miRNA read count data were normalized using the DESeq2 package (33) as described in the Materials and methods . (a) Heatmap depiction of sma-miRNA abundance (represented by Z-scores) found within EV-enriched and EV-depleted supernatant fractions after agglomerative hierarchical clustering and standardization. Each row represents a miRNA and its relative Z-score value in EV-enriched and EV-depleted fractions is displayed in the 2 columns. All sma-miRNA specifics (name, sequence, raw/normalized read counts and cluster location) are included in Supplementary file 3 . (b) sma-miRNA localization found throughout the S. mansoni karyotype (v5.2). Vertical grey bars represent the position of known sma-miRNAs available in miRBase (v.21). Vertical black bars represent the localization of all extracellular sma-miRNAs (within EV-depleted and EV-enriched fractions) newly identified in our study. Black stars above the vertical lines represent 13 sma-miRNAs found in our study that are also present in miRBase. Eighteen miRNAs localized on unmapped scaffolds (16 not mapped at all; 1 unplaced on Ch 7, 4 and 3; 5 unplaced on Ch 1) as well as 15 miRNAs not yet mapped to the current S. mansoni genome assembly were not included in this analysis. All available sma-miRNA localization coordinates are available in Supplementary file 3 .
    Nebnext Index Primer For Illumina, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Mapping nucleosome distributions following KSHV reactivation using mTSS-seq technique and mTSS-seq validation A. Experimental design for mapping nucleosome distributions following KSHV reactivation using our newly developed mTSS-seq technique. B. Validation of mTSS-seq using quantitative PCR for both on-target regions of the genome (within the sequence-capture region, two-kb region centered on a TSS) and off-target regions of the genome (not within the sequence-capture region, outside the two-region centered on a TSS). Quantitative PCR was performed for on- and off-target regions of the genome for both RHOC and ITGA4. The y-axis shows C t values. On average, C t values between the on- and off-targets of the sequence-captured libraries differ by 10.5 cycles. C. The periodic occurrence of AA/TT/AT/TA dinucleotides was calculated for all nucleosomal-sized fragments (147-148 bp) at the 0 hour time point. The x-axis represents the distance from the dyad axis. The y-axis is the frequency of AA/TT/AT/TA dinucleotides. An A/T-containing dinucleotide periodicity is seen every 10 bp at the 0 hour time point. D. Alignment of the midpoint fragments (purple) from mTSS-seq to the human genome shown in the UCSC Genome Browser ( http://genome.ucsc.edu ). Zooming in 5000X on human chromosome 2 to a six-kb window with two-kb of midpoint fragments at 0 hour time point (purple lines), along with the sequence-capture oligos and previously-published human-nucleosome distribution map for cell line GM18508 (red lines, (Gaffney et al., 2012)) at the TSSs of PUS10 and PEX13. The x-axis is genomic location. The y-axis is scaled reads per million.

    Journal: Oncotarget

    Article Title: Hierarchical regulation of the genome: global changes in nucleosome organization potentiate genome response

    doi: 10.18632/oncotarget.6841

    Figure Lengend Snippet: Mapping nucleosome distributions following KSHV reactivation using mTSS-seq technique and mTSS-seq validation A. Experimental design for mapping nucleosome distributions following KSHV reactivation using our newly developed mTSS-seq technique. B. Validation of mTSS-seq using quantitative PCR for both on-target regions of the genome (within the sequence-capture region, two-kb region centered on a TSS) and off-target regions of the genome (not within the sequence-capture region, outside the two-region centered on a TSS). Quantitative PCR was performed for on- and off-target regions of the genome for both RHOC and ITGA4. The y-axis shows C t values. On average, C t values between the on- and off-targets of the sequence-captured libraries differ by 10.5 cycles. C. The periodic occurrence of AA/TT/AT/TA dinucleotides was calculated for all nucleosomal-sized fragments (147-148 bp) at the 0 hour time point. The x-axis represents the distance from the dyad axis. The y-axis is the frequency of AA/TT/AT/TA dinucleotides. An A/T-containing dinucleotide periodicity is seen every 10 bp at the 0 hour time point. D. Alignment of the midpoint fragments (purple) from mTSS-seq to the human genome shown in the UCSC Genome Browser ( http://genome.ucsc.edu ). Zooming in 5000X on human chromosome 2 to a six-kb window with two-kb of midpoint fragments at 0 hour time point (purple lines), along with the sequence-capture oligos and previously-published human-nucleosome distribution map for cell line GM18508 (red lines, (Gaffney et al., 2012)) at the TSSs of PUS10 and PEX13. The x-axis is genomic location. The y-axis is scaled reads per million.

    Article Snippet: The universal and indexed sequences were added by PCR using 23 ul of adaptor-ligated DNA fragments, NEBNext High Fidelity 2X PCR Master Mix, index primers provided in NEBNext Multiplex (NEB #E7335, #E7500) Oligos for Illumina, and Universal PCR Primers provided in NEBNext Multiplex (NEB #E7335, #E7500) Oligos for Illumina.

    Techniques: Real-time Polymerase Chain Reaction, Sequencing

    The newly developed mTSS-Capture method combined with paired-end sequencing maps genome-wide nucleosome distribution in primary patient samples and identifies bona fide nucleosome characteristics, concordant with other human nucleosome mapping studies A . Work-flow of the mTSS-seq method. Following MNase digestion using a titration of MNase, populations of mononucleosomally protected DNA and subnucleosomal fragments are isolated, and prepared as libraries for Illumina sequencing. Solution-based sequence capture is performed using biotinylated oligos, enabling the enrichment of fragments within 2kb of each transcription start site in the human genome. Paired-end 50bp sequencing was then performed on each index. B . Alignment of the mTSS-seq midpoints to the human genome using the UCSC genome browser for LAC patient #4137 Normal tissue is shown for chr11, hg19 ( http://genome.ucsc.edu ). Zooming in twice at 100X allows for further visualization of the sequence capture oligos surrounding the TSS in a 500kb and a 5kb region showing the ATM locus. C . Averaged, normalized reads per million (y-axis) from mTSS-seq plotted as fragments (gray) and midpoints (black), centered on and surrounding 2kb of the TSS for ~22,000 open reading frames in hg19 (x-axis). DNase I-hypersensitivity (GSM736580; green) and RNA polymerase II from ChIP-seq (GSM935299; blue) data from A549 cells are shown. (D) LAC patient 4137 Normal nucleosomal midpoints (blue track) were plotted in the UCSC genome browser against the published human lymphocyte nucleosome distribution maps by Gaffney et. al. (green track) for the ZNF451 and CCDC97 loci. Sequence capture oligos and corresponding RefSeq gene models are shown for each locus. Correlations are shown for ZNF451 and CCDC87, respectively.

    Journal: Oncotarget

    Article Title: Comprehensive nucleosome mapping of the human genome in cancer progression

    doi: 10.18632/oncotarget.6811

    Figure Lengend Snippet: The newly developed mTSS-Capture method combined with paired-end sequencing maps genome-wide nucleosome distribution in primary patient samples and identifies bona fide nucleosome characteristics, concordant with other human nucleosome mapping studies A . Work-flow of the mTSS-seq method. Following MNase digestion using a titration of MNase, populations of mononucleosomally protected DNA and subnucleosomal fragments are isolated, and prepared as libraries for Illumina sequencing. Solution-based sequence capture is performed using biotinylated oligos, enabling the enrichment of fragments within 2kb of each transcription start site in the human genome. Paired-end 50bp sequencing was then performed on each index. B . Alignment of the mTSS-seq midpoints to the human genome using the UCSC genome browser for LAC patient #4137 Normal tissue is shown for chr11, hg19 ( http://genome.ucsc.edu ). Zooming in twice at 100X allows for further visualization of the sequence capture oligos surrounding the TSS in a 500kb and a 5kb region showing the ATM locus. C . Averaged, normalized reads per million (y-axis) from mTSS-seq plotted as fragments (gray) and midpoints (black), centered on and surrounding 2kb of the TSS for ~22,000 open reading frames in hg19 (x-axis). DNase I-hypersensitivity (GSM736580; green) and RNA polymerase II from ChIP-seq (GSM935299; blue) data from A549 cells are shown. (D) LAC patient 4137 Normal nucleosomal midpoints (blue track) were plotted in the UCSC genome browser against the published human lymphocyte nucleosome distribution maps by Gaffney et. al. (green track) for the ZNF451 and CCDC97 loci. Sequence capture oligos and corresponding RefSeq gene models are shown for each locus. Correlations are shown for ZNF451 and CCDC87, respectively.

    Article Snippet: Universal and indexed sequences were added through 8 cycles of PCR, using NEBNext® Multiplex Oligos for Illumina® (Index Primers Set 1, NEB #E7335S/L).

    Techniques: Sequencing, Genome Wide, Titration, Isolation, Chromatin Immunoprecipitation

    Schistosomula excrete/secrete extracellular miRNAs. Supernatant from 72 h in vitro cultured schistosomula was separated into EV-enriched and EV-depleted fractions (n=3) by preparatory ultracentrifugation. Total RNA from each fraction was extracted using the miRNeasy kit (Qiagen) and prepared for RNA-seq using the NEBNext Small RNA (for Illumina HiSeq sequencing) kit. To identify sma-miRNAs in our samples, the miRDeep2 package was utilized (29). Only miRNAs with at least 10 reads in 2 out of the 3 biological replicates (in at least one of the fractions, EV-enriched OR EV-depleted) were considered in the study. A total of 205 putative sma-miRNAs passed this criterion. sma-miRNA read count data were normalized using the DESeq2 package (33) as described in the Materials and methods . (a) Heatmap depiction of sma-miRNA abundance (represented by Z-scores) found within EV-enriched and EV-depleted supernatant fractions after agglomerative hierarchical clustering and standardization. Each row represents a miRNA and its relative Z-score value in EV-enriched and EV-depleted fractions is displayed in the 2 columns. All sma-miRNA specifics (name, sequence, raw/normalized read counts and cluster location) are included in Supplementary file 3 . (b) sma-miRNA localization found throughout the S. mansoni karyotype (v5.2). Vertical grey bars represent the position of known sma-miRNAs available in miRBase (v.21). Vertical black bars represent the localization of all extracellular sma-miRNAs (within EV-depleted and EV-enriched fractions) newly identified in our study. Black stars above the vertical lines represent 13 sma-miRNAs found in our study that are also present in miRBase. Eighteen miRNAs localized on unmapped scaffolds (16 not mapped at all; 1 unplaced on Ch 7, 4 and 3; 5 unplaced on Ch 1) as well as 15 miRNAs not yet mapped to the current S. mansoni genome assembly were not included in this analysis. All available sma-miRNA localization coordinates are available in Supplementary file 3 .

    Journal: Journal of Extracellular Vesicles

    Article Title: Protein and small non-coding RNA-enriched extracellular vesicles are released by the pathogenic blood fluke Schistosoma mansoni

    doi: 10.3402/jev.v4.28665

    Figure Lengend Snippet: Schistosomula excrete/secrete extracellular miRNAs. Supernatant from 72 h in vitro cultured schistosomula was separated into EV-enriched and EV-depleted fractions (n=3) by preparatory ultracentrifugation. Total RNA from each fraction was extracted using the miRNeasy kit (Qiagen) and prepared for RNA-seq using the NEBNext Small RNA (for Illumina HiSeq sequencing) kit. To identify sma-miRNAs in our samples, the miRDeep2 package was utilized (29). Only miRNAs with at least 10 reads in 2 out of the 3 biological replicates (in at least one of the fractions, EV-enriched OR EV-depleted) were considered in the study. A total of 205 putative sma-miRNAs passed this criterion. sma-miRNA read count data were normalized using the DESeq2 package (33) as described in the Materials and methods . (a) Heatmap depiction of sma-miRNA abundance (represented by Z-scores) found within EV-enriched and EV-depleted supernatant fractions after agglomerative hierarchical clustering and standardization. Each row represents a miRNA and its relative Z-score value in EV-enriched and EV-depleted fractions is displayed in the 2 columns. All sma-miRNA specifics (name, sequence, raw/normalized read counts and cluster location) are included in Supplementary file 3 . (b) sma-miRNA localization found throughout the S. mansoni karyotype (v5.2). Vertical grey bars represent the position of known sma-miRNAs available in miRBase (v.21). Vertical black bars represent the localization of all extracellular sma-miRNAs (within EV-depleted and EV-enriched fractions) newly identified in our study. Black stars above the vertical lines represent 13 sma-miRNAs found in our study that are also present in miRBase. Eighteen miRNAs localized on unmapped scaffolds (16 not mapped at all; 1 unplaced on Ch 7, 4 and 3; 5 unplaced on Ch 1) as well as 15 miRNAs not yet mapped to the current S. mansoni genome assembly were not included in this analysis. All available sma-miRNA localization coordinates are available in Supplementary file 3 .

    Article Snippet: Small RNA concentrations ranged from 60 pg/ml (EV-enriched fraction) to 750 pg/ml (EV-depleted fraction) as determined by the Bioanalyzer. sncRNA libraries were prepared using the NEBNext Multiplex Small RNA Library Prep Set and NEBNext index primer for Illumina (New England Biolabs, Ipswich, MA, USA), according to the manufacturer's protocol.

    Techniques: In Vitro, Cell Culture, RNA Sequencing Assay, Sequencing