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    Illumina Inc illumina miseq platform
    Illustration of the process workflow. 1.) Cells are diluted in alginate polymer to a concentration of approximately 10 5 cells per microliter, resulting in a 10% occupancy rate in 100 μm microspheres. 2.) Cells are lysed using heat, and the bulk microsphere solids are mixed with reagents for a 2-step whole genome amplification reaction. 3.) After amplification, microspheres are diluted to extinction in a 384 well plate, and scanned for presence of single microspheres that fluoresce with PicoGreen DNA stain. 4.) An isolated microsphere is transferred to a fresh tube and the DNA products are recovered by dissolving the alginate matrix. These amplified products are submitted to further rounds of amplification to add sequencing adapters. 5.) The products are prepared for high throughput sequencing using the <t>Illumina</t> <t>MiSeq</t> platform. Intermediate steps include fluorescence microscopy and quantitative PCR for quality control.
    Illumina Miseq Platform, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 99/100, based on 42244 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/illumina miseq platform/product/Illumina Inc
    Average 99 stars, based on 42244 article reviews
    Price from $9.99 to $1999.99
    illumina miseq platform - by Bioz Stars, 2020-07
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    88
    Illumina Inc miseq ngs platform
    Illustration of the process workflow. 1.) Cells are diluted in alginate polymer to a concentration of approximately 10 5 cells per microliter, resulting in a 10% occupancy rate in 100 μm microspheres. 2.) Cells are lysed using heat, and the bulk microsphere solids are mixed with reagents for a 2-step whole genome amplification reaction. 3.) After amplification, microspheres are diluted to extinction in a 384 well plate, and scanned for presence of single microspheres that fluoresce with PicoGreen DNA stain. 4.) An isolated microsphere is transferred to a fresh tube and the DNA products are recovered by dissolving the alginate matrix. These amplified products are submitted to further rounds of amplification to add sequencing adapters. 5.) The products are prepared for high throughput sequencing using the <t>Illumina</t> <t>MiSeq</t> platform. Intermediate steps include fluorescence microscopy and quantitative PCR for quality control.
    Miseq Ngs Platform, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 88/100, based on 148 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/miseq ngs platform/product/Illumina Inc
    Average 88 stars, based on 148 article reviews
    Price from $9.99 to $1999.99
    miseq ngs platform - by Bioz Stars, 2020-07
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    90
    Illumina Inc miseq 250 platform
    Illustration of the process workflow. 1.) Cells are diluted in alginate polymer to a concentration of approximately 10 5 cells per microliter, resulting in a 10% occupancy rate in 100 μm microspheres. 2.) Cells are lysed using heat, and the bulk microsphere solids are mixed with reagents for a 2-step whole genome amplification reaction. 3.) After amplification, microspheres are diluted to extinction in a 384 well plate, and scanned for presence of single microspheres that fluoresce with PicoGreen DNA stain. 4.) An isolated microsphere is transferred to a fresh tube and the DNA products are recovered by dissolving the alginate matrix. These amplified products are submitted to further rounds of amplification to add sequencing adapters. 5.) The products are prepared for high throughput sequencing using the <t>Illumina</t> <t>MiSeq</t> platform. Intermediate steps include fluorescence microscopy and quantitative PCR for quality control.
    Miseq 250 Platform, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 90/100, based on 20 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 90 stars, based on 20 article reviews
    Price from $9.99 to $1999.99
    miseq 250 platform - by Bioz Stars, 2020-07
    90/100 stars
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    90
    Illumina Inc miseq pe250 platform
    Illustration of the process workflow. 1.) Cells are diluted in alginate polymer to a concentration of approximately 10 5 cells per microliter, resulting in a 10% occupancy rate in 100 μm microspheres. 2.) Cells are lysed using heat, and the bulk microsphere solids are mixed with reagents for a 2-step whole genome amplification reaction. 3.) After amplification, microspheres are diluted to extinction in a 384 well plate, and scanned for presence of single microspheres that fluoresce with PicoGreen DNA stain. 4.) An isolated microsphere is transferred to a fresh tube and the DNA products are recovered by dissolving the alginate matrix. These amplified products are submitted to further rounds of amplification to add sequencing adapters. 5.) The products are prepared for high throughput sequencing using the <t>Illumina</t> <t>MiSeq</t> platform. Intermediate steps include fluorescence microscopy and quantitative PCR for quality control.
    Miseq Pe250 Platform, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 90/100, based on 232 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/miseq pe250 platform/product/Illumina Inc
    Average 90 stars, based on 232 article reviews
    Price from $9.99 to $1999.99
    miseq pe250 platform - by Bioz Stars, 2020-07
    90/100 stars
      Buy from Supplier

    Image Search Results


    Illustration of the process workflow. 1.) Cells are diluted in alginate polymer to a concentration of approximately 10 5 cells per microliter, resulting in a 10% occupancy rate in 100 μm microspheres. 2.) Cells are lysed using heat, and the bulk microsphere solids are mixed with reagents for a 2-step whole genome amplification reaction. 3.) After amplification, microspheres are diluted to extinction in a 384 well plate, and scanned for presence of single microspheres that fluoresce with PicoGreen DNA stain. 4.) An isolated microsphere is transferred to a fresh tube and the DNA products are recovered by dissolving the alginate matrix. These amplified products are submitted to further rounds of amplification to add sequencing adapters. 5.) The products are prepared for high throughput sequencing using the Illumina MiSeq platform. Intermediate steps include fluorescence microscopy and quantitative PCR for quality control.

    Journal: PLoS ONE

    Article Title: A Simple Method for Encapsulating Single Cells in Alginate Microspheres Allows for Direct PCR and Whole Genome Amplification

    doi: 10.1371/journal.pone.0117738

    Figure Lengend Snippet: Illustration of the process workflow. 1.) Cells are diluted in alginate polymer to a concentration of approximately 10 5 cells per microliter, resulting in a 10% occupancy rate in 100 μm microspheres. 2.) Cells are lysed using heat, and the bulk microsphere solids are mixed with reagents for a 2-step whole genome amplification reaction. 3.) After amplification, microspheres are diluted to extinction in a 384 well plate, and scanned for presence of single microspheres that fluoresce with PicoGreen DNA stain. 4.) An isolated microsphere is transferred to a fresh tube and the DNA products are recovered by dissolving the alginate matrix. These amplified products are submitted to further rounds of amplification to add sequencing adapters. 5.) The products are prepared for high throughput sequencing using the Illumina MiSeq platform. Intermediate steps include fluorescence microscopy and quantitative PCR for quality control.

    Article Snippet: This quantity of DNA is six orders of magnitude less material than required for the current Illumina MiSeq platform.

    Techniques: Concentration Assay, Whole Genome Amplification, Amplification, Staining, Isolation, Sequencing, Next-Generation Sequencing, Fluorescence, Microscopy, Real-time Polymerase Chain Reaction

    Schematic overview of microbiome bioinformatic analysis workflow. The hypervariable V4 region of 16S rDNA from tick samples was sequenced and split by barcode with Illumina MiSeq. Resulting paired-end reads were joined and the primer region was removed. Reads were filtered by amplicon length and aligned to SILVA as the reference database. After removal of chimeras, reads were clustered into operational taxonomic units (OTU) and taxonomically classified. Finally, an OTU-table was created and results were visualized

    Journal: Parasites & Vectors

    Article Title: Combination of microbiome analysis and serodiagnostics to assess the risk of pathogen transmission by ticks to humans and animals in central Germany

    doi: 10.1186/s13071-018-3240-7

    Figure Lengend Snippet: Schematic overview of microbiome bioinformatic analysis workflow. The hypervariable V4 region of 16S rDNA from tick samples was sequenced and split by barcode with Illumina MiSeq. Resulting paired-end reads were joined and the primer region was removed. Reads were filtered by amplicon length and aligned to SILVA as the reference database. After removal of chimeras, reads were clustered into operational taxonomic units (OTU) and taxonomically classified. Finally, an OTU-table was created and results were visualized

    Article Snippet: All samples were diluted to the same molarity, pooled together, spiked with an internal control (15% PhiX) and paired-end sequenced on the MiSeq Illumina platform using a flow cell with V2 chemistry (500 cycles).

    Techniques: Amplification

    —Examples of sequence alignments between Bemisia tabaci “Peru” MEAM1 mtDNA COI haplotype gene region, published “MEAM2” mtDNA COI gene region, and NGS candidate NUMT sequences identified from KX234913, KY951449, and the KY951452 individuals. ( A ) C-terminal region of a Peruvian MEAM1 B. tabaci (KY951450) mtDNA COI gene region showing putative stop codon (black shaded “*” symbol), as well as the B. tabaci “MEAM2” haplotype from Japan (AJ550177), and examples NGS candidate NUMT sequences from the Peruvian B. tabaci MEAM1 (KX234914) with matching SNPs (indicated by red boxes) that matched the Japan MEAM2 haplotype (AJ550177). Deletion of a “T” base (indicated by red triangle) resulted in a frameshift mutation and the loss of the putative mtDNA COI gene stop codon. ( B ) Internal stop codons (at positions 904S906) detected in candidate NUMT sequences (KY951452_NUMT-01, KY951452_NUMT-02) from the Peruvian MEAM1 individual (KY951452) MiSeq generated DNA fragments when compared with the Peruvian B. tabaci MEAM1 (KX234914) mtDNA COI gene. Stop codons detected in NUMT sequences were the result of a single nucleotide base change at position 906 from a “T” to an “A.” Candidate NUMT sequences were also compared with the Peruvian MEAM2 haplotype (KY951454) obtained via PCR and Sanger sequencing of the same MEAM1 individual (KY951452). Nucleotide positions based on the mtDNA COI gene are provided. Amino acid translation based on the invertebrate mitochondrial genetic codes (Translational Table_5). Significant changes between amino acids are highlighted.

    Journal: Genome Biology and Evolution

    Article Title: The Trouble with MEAM2: Implications of Pseudogenes on Species Delimitation in the Globally Invasive Bemisia tabaci (Hemiptera: Aleyrodidae) Cryptic Species Complex

    doi: 10.1093/gbe/evx173

    Figure Lengend Snippet: —Examples of sequence alignments between Bemisia tabaci “Peru” MEAM1 mtDNA COI haplotype gene region, published “MEAM2” mtDNA COI gene region, and NGS candidate NUMT sequences identified from KX234913, KY951449, and the KY951452 individuals. ( A ) C-terminal region of a Peruvian MEAM1 B. tabaci (KY951450) mtDNA COI gene region showing putative stop codon (black shaded “*” symbol), as well as the B. tabaci “MEAM2” haplotype from Japan (AJ550177), and examples NGS candidate NUMT sequences from the Peruvian B. tabaci MEAM1 (KX234914) with matching SNPs (indicated by red boxes) that matched the Japan MEAM2 haplotype (AJ550177). Deletion of a “T” base (indicated by red triangle) resulted in a frameshift mutation and the loss of the putative mtDNA COI gene stop codon. ( B ) Internal stop codons (at positions 904S906) detected in candidate NUMT sequences (KY951452_NUMT-01, KY951452_NUMT-02) from the Peruvian MEAM1 individual (KY951452) MiSeq generated DNA fragments when compared with the Peruvian B. tabaci MEAM1 (KX234914) mtDNA COI gene. Stop codons detected in NUMT sequences were the result of a single nucleotide base change at position 906 from a “T” to an “A.” Candidate NUMT sequences were also compared with the Peruvian MEAM2 haplotype (KY951454) obtained via PCR and Sanger sequencing of the same MEAM1 individual (KY951452). Nucleotide positions based on the mtDNA COI gene are provided. Amino acid translation based on the invertebrate mitochondrial genetic codes (Translational Table_5). Significant changes between amino acids are highlighted.

    Article Snippet: We identified low copy genome fragments through the Illumina MiSeq sequencing platform in MEAM1 individuals that matched unique MEAM2 SNPs ( ).

    Techniques: Sequencing, Next-Generation Sequencing, Mutagenesis, Generated, Polymerase Chain Reaction

    Methylation analysis comparison of the 35S-eGFP+hp time course experiment. (A) Methylation analysis was performed after the samples were sequenced using the Ion Torrent. A minimum length of 395 bp was selected with Phred scores of Q12 for 2 and 3 dpi, and Q22 for the remaining time points. (B) Methylation analysis on the same samples sequenced using the Illumina MiSeq tagmentation approach. A minimum length of 250 bp was selected with Phred scores of Q30 for all samples. n = number of reads used from one infiltrated leaf to build the profile after quality and length filtering.

    Journal: Frontiers in Plant Science

    Article Title: The Rapid Methylation of T-DNAs Upon Agrobacterium Inoculation in Plant Leaves

    doi: 10.3389/fpls.2019.00312

    Figure Lengend Snippet: Methylation analysis comparison of the 35S-eGFP+hp time course experiment. (A) Methylation analysis was performed after the samples were sequenced using the Ion Torrent. A minimum length of 395 bp was selected with Phred scores of Q12 for 2 and 3 dpi, and Q22 for the remaining time points. (B) Methylation analysis on the same samples sequenced using the Illumina MiSeq tagmentation approach. A minimum length of 250 bp was selected with Phred scores of Q30 for all samples. n = number of reads used from one infiltrated leaf to build the profile after quality and length filtering.

    Article Snippet: Comparisons Between Ion Torrent and Illumina MiSeq As more T-DNA amplicon sequence reads are able to be cost-effectively assessed by NGS rather than Sanger sequencing, a time course of the methylation profile compared only the Ion Torrent and Illumina MiSeq sequencing platforms.

    Techniques: Methylation

    Methylation analysis comparison in the time course of the AtEF1α-A4 promoter with (left panel) or without the 5′ UTR intron (right panel) regulating the expression of eGFP . Percentage cytosine methylation is presented on the y-axis and individual cytosine positions in the 546 bp region indicated by the black bar on the x-axis. Sequencing was carried out on the Illumina MiSeq with the fusion protocol, n = three biological replicates. General linear model (two-way ANOVA) followed by the Tukey test was used to compare all cytosine contexts between the two treatments at each time point. Statistically significant differences, ns not significant, ∗ P

    Journal: Frontiers in Plant Science

    Article Title: The Rapid Methylation of T-DNAs Upon Agrobacterium Inoculation in Plant Leaves

    doi: 10.3389/fpls.2019.00312

    Figure Lengend Snippet: Methylation analysis comparison in the time course of the AtEF1α-A4 promoter with (left panel) or without the 5′ UTR intron (right panel) regulating the expression of eGFP . Percentage cytosine methylation is presented on the y-axis and individual cytosine positions in the 546 bp region indicated by the black bar on the x-axis. Sequencing was carried out on the Illumina MiSeq with the fusion protocol, n = three biological replicates. General linear model (two-way ANOVA) followed by the Tukey test was used to compare all cytosine contexts between the two treatments at each time point. Statistically significant differences, ns not significant, ∗ P

    Article Snippet: Comparisons Between Ion Torrent and Illumina MiSeq As more T-DNA amplicon sequence reads are able to be cost-effectively assessed by NGS rather than Sanger sequencing, a time course of the methylation profile compared only the Ion Torrent and Illumina MiSeq sequencing platforms.

    Techniques: Methylation, Expressing, Sequencing

    Sequencing platform comparison between Sanger sequencing, Ion Torrent PGM and Illumina MiSeq using the tagamntation approach. The percentage of methylated cytosines (% mCs) is presented for the 14 dpi 35S-eGFP+hp sample, n = number of reads used from one infiltrated leaf to build the profile after quality and length filtering.

    Journal: Frontiers in Plant Science

    Article Title: The Rapid Methylation of T-DNAs Upon Agrobacterium Inoculation in Plant Leaves

    doi: 10.3389/fpls.2019.00312

    Figure Lengend Snippet: Sequencing platform comparison between Sanger sequencing, Ion Torrent PGM and Illumina MiSeq using the tagamntation approach. The percentage of methylated cytosines (% mCs) is presented for the 14 dpi 35S-eGFP+hp sample, n = number of reads used from one infiltrated leaf to build the profile after quality and length filtering.

    Article Snippet: Comparisons Between Ion Torrent and Illumina MiSeq As more T-DNA amplicon sequence reads are able to be cost-effectively assessed by NGS rather than Sanger sequencing, a time course of the methylation profile compared only the Ion Torrent and Illumina MiSeq sequencing platforms.

    Techniques: Sequencing, Methylation

    Methylation analysis comparison in the time course of the 35S-eGFP transgene without a hairpin (left panel) or with a hairpin (right panel) targeting the eGFP transgene (shown in purple). Percentage cytosine methylation is presented on the y-axis and individual cytosine positions in the 418 bp region indicated by the black bar on the x-axis. Background highlights: blue: promoter, white: 5′ UTR and green: coding sequence (CDS). Sequencing was carried out on the Illumina MiSeq with the fusion protocol, n = three biological replicates. General linear model (two-way ANOVA) followed by the Tukey test was used to compare all cytosine contexts between the two treatments at each time point. Statistically significant differences, ns not significant, ∗ P

    Journal: Frontiers in Plant Science

    Article Title: The Rapid Methylation of T-DNAs Upon Agrobacterium Inoculation in Plant Leaves

    doi: 10.3389/fpls.2019.00312

    Figure Lengend Snippet: Methylation analysis comparison in the time course of the 35S-eGFP transgene without a hairpin (left panel) or with a hairpin (right panel) targeting the eGFP transgene (shown in purple). Percentage cytosine methylation is presented on the y-axis and individual cytosine positions in the 418 bp region indicated by the black bar on the x-axis. Background highlights: blue: promoter, white: 5′ UTR and green: coding sequence (CDS). Sequencing was carried out on the Illumina MiSeq with the fusion protocol, n = three biological replicates. General linear model (two-way ANOVA) followed by the Tukey test was used to compare all cytosine contexts between the two treatments at each time point. Statistically significant differences, ns not significant, ∗ P

    Article Snippet: Comparisons Between Ion Torrent and Illumina MiSeq As more T-DNA amplicon sequence reads are able to be cost-effectively assessed by NGS rather than Sanger sequencing, a time course of the methylation profile compared only the Ion Torrent and Illumina MiSeq sequencing platforms.

    Techniques: Methylation, Sequencing

    (A) The percentage of methylated cytosines (% mCs) is presented for the 14 dpi 35S-eGFP+hp sample sequenced on the Illumina MiSeq with the fusion protocol. Comparisons are amongst the DNA polymerases (KAPA2G and NEB EpiMark) and extension temperature used during the bisulfite PCR; n = number of reads used from one infiltrated leaf to build the profile after quality and length filtering. (B) Gel electrophoresis showing amplification intensity of the same sample used to generate the methylation profile in (A) ; NTC, no template control. 100 bp DNA Ladder (GeneRuler).

    Journal: Frontiers in Plant Science

    Article Title: The Rapid Methylation of T-DNAs Upon Agrobacterium Inoculation in Plant Leaves

    doi: 10.3389/fpls.2019.00312

    Figure Lengend Snippet: (A) The percentage of methylated cytosines (% mCs) is presented for the 14 dpi 35S-eGFP+hp sample sequenced on the Illumina MiSeq with the fusion protocol. Comparisons are amongst the DNA polymerases (KAPA2G and NEB EpiMark) and extension temperature used during the bisulfite PCR; n = number of reads used from one infiltrated leaf to build the profile after quality and length filtering. (B) Gel electrophoresis showing amplification intensity of the same sample used to generate the methylation profile in (A) ; NTC, no template control. 100 bp DNA Ladder (GeneRuler).

    Article Snippet: Comparisons Between Ion Torrent and Illumina MiSeq As more T-DNA amplicon sequence reads are able to be cost-effectively assessed by NGS rather than Sanger sequencing, a time course of the methylation profile compared only the Ion Torrent and Illumina MiSeq sequencing platforms.

    Techniques: Methylation, Polymerase Chain Reaction, Nucleic Acid Electrophoresis, Amplification

    High-throughput workflow for the preparation of plasmid DNA libraries for NGS. Plasmid DNA samples are isolated from bacteria cells using a high-throughput plasmid isolation method on the CyBio ® FeliX robot. All steps performed using the FeliX platform are highlighted with a grey outline. The isolated plasmid samples are tested for the presence of genomic DNA (gDNA), prior to library preparation, using the Labcyte Echo ® . All steps performed using the Labcyte Echo ® are highlighted with a shaded grey box. If samples are free from gDNA, they are diluted to 0.4 ng/μl in H 2 O. If gDNA is detected, the samples are sonicated prior to re-testing in the gDNA QC assay. Using reagents from the Nextera XT kit, a tagmentation reaction is performed on all samples under the optimised conditions, followed by neutralization of the reaction. Unique combinations of index primers are added to all samples via 12 PCR cycles, followed by magnetic bead purification of the PCR products. The concentration of the purified dsDNA is then determined using the PicoGreen ® reagent assay and the libraries are pooled to give a final concentration of 4–10 nM, in a minimum volume of 15 μl. The average fragment size of the pooled libraries is measured using the Fragment Analyzer before being sequenced on the Illumina ® MiSeq system.

    Journal: Synthetic and Systems Biotechnology

    Article Title: Miniaturisation of high-throughput plasmid DNA library preparation for next-generation sequencing using multifactorial optimisation

    doi: 10.1016/j.synbio.2019.01.002

    Figure Lengend Snippet: High-throughput workflow for the preparation of plasmid DNA libraries for NGS. Plasmid DNA samples are isolated from bacteria cells using a high-throughput plasmid isolation method on the CyBio ® FeliX robot. All steps performed using the FeliX platform are highlighted with a grey outline. The isolated plasmid samples are tested for the presence of genomic DNA (gDNA), prior to library preparation, using the Labcyte Echo ® . All steps performed using the Labcyte Echo ® are highlighted with a shaded grey box. If samples are free from gDNA, they are diluted to 0.4 ng/μl in H 2 O. If gDNA is detected, the samples are sonicated prior to re-testing in the gDNA QC assay. Using reagents from the Nextera XT kit, a tagmentation reaction is performed on all samples under the optimised conditions, followed by neutralization of the reaction. Unique combinations of index primers are added to all samples via 12 PCR cycles, followed by magnetic bead purification of the PCR products. The concentration of the purified dsDNA is then determined using the PicoGreen ® reagent assay and the libraries are pooled to give a final concentration of 4–10 nM, in a minimum volume of 15 μl. The average fragment size of the pooled libraries is measured using the Fragment Analyzer before being sequenced on the Illumina ® MiSeq system.

    Article Snippet: 3.3 Optimisation of plasmid DNA library preparation for NGS using Design of Experiments For sequencing of plasmid DNA using the Illumina ® MiSeq platform, a library of fragments with a mean size of between 200 and 400 bp, each with a concentration of 0.5–5 ng/μl is required.

    Techniques: High Throughput Screening Assay, Plasmid Preparation, Next-Generation Sequencing, Isolation, Sonication, Neutralization, Polymerase Chain Reaction, Purification, Concentration Assay

    Next generation sequencing of plasmid DNA libraries, prepared using a miniaturised method with the Nextera XT library preparation kit. 96 plasmid DNA libraries were prepared for NGS with the Nextera XT library preparation kit, using optimised conditions (12.5 min incubation, 50 nl sample, 1.8x magnetic bead solution). ( A ) After purification, the samples were quantified in the PicoGreen ® dsDNA quantification assay. The data show that 75/92 samples have a concentration within the desired range (0.5–5 ng/μl), with an average concentration of 1.1 ng/μl. These samples were pooled, with a final concentration of 6.64 nM and run on the Fragment Analyzer ( B ) and ( C ). All fragments in the pooled libraries are of the desired size (200–400 bp). ( D ) The pooled library was sequenced on the Illumina ® MiSeq system (2 × 150 method). The mean sequence quality (Phred) scores are plotted for each sample. For all samples, the sequence quality (Phred) score was > 30 for more than 85% of the base pairs, indicating that all samples passed the QC criteria.

    Journal: Synthetic and Systems Biotechnology

    Article Title: Miniaturisation of high-throughput plasmid DNA library preparation for next-generation sequencing using multifactorial optimisation

    doi: 10.1016/j.synbio.2019.01.002

    Figure Lengend Snippet: Next generation sequencing of plasmid DNA libraries, prepared using a miniaturised method with the Nextera XT library preparation kit. 96 plasmid DNA libraries were prepared for NGS with the Nextera XT library preparation kit, using optimised conditions (12.5 min incubation, 50 nl sample, 1.8x magnetic bead solution). ( A ) After purification, the samples were quantified in the PicoGreen ® dsDNA quantification assay. The data show that 75/92 samples have a concentration within the desired range (0.5–5 ng/μl), with an average concentration of 1.1 ng/μl. These samples were pooled, with a final concentration of 6.64 nM and run on the Fragment Analyzer ( B ) and ( C ). All fragments in the pooled libraries are of the desired size (200–400 bp). ( D ) The pooled library was sequenced on the Illumina ® MiSeq system (2 × 150 method). The mean sequence quality (Phred) scores are plotted for each sample. For all samples, the sequence quality (Phred) score was > 30 for more than 85% of the base pairs, indicating that all samples passed the QC criteria.

    Article Snippet: 3.3 Optimisation of plasmid DNA library preparation for NGS using Design of Experiments For sequencing of plasmid DNA using the Illumina ® MiSeq platform, a library of fragments with a mean size of between 200 and 400 bp, each with a concentration of 0.5–5 ng/μl is required.

    Techniques: Next-Generation Sequencing, Plasmid Preparation, Incubation, Purification, Concentration Assay, Sequencing

    Workflow using the 16S Library Preparation Protocol in Illumina MiSeq platform.

    Journal: Frontiers in Immunology

    Article Title: Intestinal Dysbiosis in Autoimmune Diabetes Is Correlated With Poor Glycemic Control and Increased Interleukin-6: A Pilot Study

    doi: 10.3389/fimmu.2018.01689

    Figure Lengend Snippet: Workflow using the 16S Library Preparation Protocol in Illumina MiSeq platform.

    Article Snippet: The sequencing was performed by using an Illumina MiSeq platform system (MiSeq Desktop Sequencer).

    Techniques:

    The continuity of the RadMap-based assemblies. The A. thaliana chromosomes are painted with assembled contigs. Alternating shades indicate adjacent contigs, and each vertical transition from gray to black represents a contig boundary or alignment breakpoint. The left half of each chromosome shows the input assembly of (A) 25× MiSeq PE300 data set, (B) 5× PacBio-5 kb data set, and (C) 5× PacBio-14 kb data set, while the right half shows the corresponding RadMap-based assembly. The RadMap-based assemblies are considerably more continuous, with 15-, 6-, and 7-fold improvement of N50 and 12-, 12-, and 5-fold improvement of N90. Chr, chromosome.

    Journal: Genetics

    Article Title: Whole-Genome Restriction Mapping by “Subhaploid”-Based RAD Sequencing: An Efficient and Flexible Approach for Physical Mapping and Genome Scaffolding

    doi: 10.1534/genetics.117.200303

    Figure Lengend Snippet: The continuity of the RadMap-based assemblies. The A. thaliana chromosomes are painted with assembled contigs. Alternating shades indicate adjacent contigs, and each vertical transition from gray to black represents a contig boundary or alignment breakpoint. The left half of each chromosome shows the input assembly of (A) 25× MiSeq PE300 data set, (B) 5× PacBio-5 kb data set, and (C) 5× PacBio-14 kb data set, while the right half shows the corresponding RadMap-based assembly. The RadMap-based assemblies are considerably more continuous, with 15-, 6-, and 7-fold improvement of N50 and 12-, 12-, and 5-fold improvement of N90. Chr, chromosome.

    Article Snippet: One paired-end DNA library with insert size of ∼500–550 bp was constructed by following the Illumina standard DNA library preparation protocol and was then sequenced using the Illumina MiSeq PE300 platform.

    Techniques:

    Examples of RadMap-linked contigs. (A) Overview and (B) zoomed-in detail of one genomic region located on the chromosome 1 (1.45–2.40 Mb) of A. thaliana , which consists of 20 contigs generated from 25× MiSeq PE300 data set, with Bsa XI tag or contig orders highly consistent with the reference genome. Chr, chromosome; Ctg, contig.

    Journal: Genetics

    Article Title: Whole-Genome Restriction Mapping by “Subhaploid”-Based RAD Sequencing: An Efficient and Flexible Approach for Physical Mapping and Genome Scaffolding

    doi: 10.1534/genetics.117.200303

    Figure Lengend Snippet: Examples of RadMap-linked contigs. (A) Overview and (B) zoomed-in detail of one genomic region located on the chromosome 1 (1.45–2.40 Mb) of A. thaliana , which consists of 20 contigs generated from 25× MiSeq PE300 data set, with Bsa XI tag or contig orders highly consistent with the reference genome. Chr, chromosome; Ctg, contig.

    Article Snippet: One paired-end DNA library with insert size of ∼500–550 bp was constructed by following the Illumina standard DNA library preparation protocol and was then sequenced using the Illumina MiSeq PE300 platform.

    Techniques: Generated, CTG Assay

    RadMap scaffolding of different WGS assemblies of A. thaliana . (A) Overview of three RadMap-based assemblies, with 15.1-, 5.7-, and 6.6-fold improvement of assembly contiguity. From inner to outer rings are genome coordinates, Bsa XI sites with between-site distances over 40 kb, and RadMap scaffolding of three WGS assemblies generated based on Illumina MiSeq PE300, PacBio-5 kb, and PacBio-14 kb data sets ( Table 3 ), respectively. The junctions between the red and green bands for the outermost three rings represent the gaps in the assembled genome, and most gaps result from genomic regions containing very sparse Bsa XI sites (between-site distances > 40 kb). (B) Dot-plot comparison of the RadMap-based assemblies and the reference genome (five chromosomes), showing high accuracy of contig linkage with Kendall’s statistic > 0.98 ( Table 3 ). One red ● represents one Bsa XI tag. Ctg, contig; Scaf, scaffold.

    Journal: Genetics

    Article Title: Whole-Genome Restriction Mapping by “Subhaploid”-Based RAD Sequencing: An Efficient and Flexible Approach for Physical Mapping and Genome Scaffolding

    doi: 10.1534/genetics.117.200303

    Figure Lengend Snippet: RadMap scaffolding of different WGS assemblies of A. thaliana . (A) Overview of three RadMap-based assemblies, with 15.1-, 5.7-, and 6.6-fold improvement of assembly contiguity. From inner to outer rings are genome coordinates, Bsa XI sites with between-site distances over 40 kb, and RadMap scaffolding of three WGS assemblies generated based on Illumina MiSeq PE300, PacBio-5 kb, and PacBio-14 kb data sets ( Table 3 ), respectively. The junctions between the red and green bands for the outermost three rings represent the gaps in the assembled genome, and most gaps result from genomic regions containing very sparse Bsa XI sites (between-site distances > 40 kb). (B) Dot-plot comparison of the RadMap-based assemblies and the reference genome (five chromosomes), showing high accuracy of contig linkage with Kendall’s statistic > 0.98 ( Table 3 ). One red ● represents one Bsa XI tag. Ctg, contig; Scaf, scaffold.

    Article Snippet: One paired-end DNA library with insert size of ∼500–550 bp was constructed by following the Illumina standard DNA library preparation protocol and was then sequenced using the Illumina MiSeq PE300 platform.

    Techniques: Scaffolding, Generated, CTG Assay

    Gap-size estimation. (A) The relationship between map distance and true physical distance. The inter-contig map distances are obtained from the RadMap assembly generated using the MiSeq PE300 data set and the corresponding true physical distances are determined according to the reference genome. The map distances range from 0 to 0.5 and are split into 50 bins. The red line refers to the average physical distance of pairs of markers for each bin, and the cyan region denotes the corresponding SE region. Note only the pairs of markers with physical distance no longer than 50-kb apart are included here. (B) Comparison of the true and predicted inter-contig gap sizes. The dashed line indicates the linear least squares fit of y = 0.7752 x + 2496, with the Pearson correlation r of 0.75.

    Journal: Genetics

    Article Title: Whole-Genome Restriction Mapping by “Subhaploid”-Based RAD Sequencing: An Efficient and Flexible Approach for Physical Mapping and Genome Scaffolding

    doi: 10.1534/genetics.117.200303

    Figure Lengend Snippet: Gap-size estimation. (A) The relationship between map distance and true physical distance. The inter-contig map distances are obtained from the RadMap assembly generated using the MiSeq PE300 data set and the corresponding true physical distances are determined according to the reference genome. The map distances range from 0 to 0.5 and are split into 50 bins. The red line refers to the average physical distance of pairs of markers for each bin, and the cyan region denotes the corresponding SE region. Note only the pairs of markers with physical distance no longer than 50-kb apart are included here. (B) Comparison of the true and predicted inter-contig gap sizes. The dashed line indicates the linear least squares fit of y = 0.7752 x + 2496, with the Pearson correlation r of 0.75.

    Article Snippet: One paired-end DNA library with insert size of ∼500–550 bp was constructed by following the Illumina standard DNA library preparation protocol and was then sequenced using the Illumina MiSeq PE300 platform.

    Techniques: Generated

    Effect of the GC content and ERCC length on the estimation of the ERCC cDNA abundance with the ONT MinION platform. The figures present deviations of the ERCC expression level estimates with the ONT MinION platform from the Ambion RNA molecular counts ( A,C ) or from the Illumina HiSeq 2500/MiSeq estimated cDNA abundance ( B,D ) as a function of the GC content ( A,B ) and the ERCC length ( C,D ). We plot the log2 ratio of observed (ONT MinION) to expected (Ambion, Illumina) read counts for the ERCC spike-ins (y-axis, log) for each of the samples relative to their length or GC content (x-axis). Due to the variable sequencing depth from each ERCC MinION experiment, each point is the average value from different MinION flow cell runs if at least 5 reads have been detected for this point in the corresponding MinION runs. The points are colored differently based on the number of flow cell runs in which they were detected (red, green, cyan, purple correspond to values derived from one, two, three or four flow cell runs respectively). The standard deviation is also presented for points with values from two or more MinION runs.

    Journal: Scientific Reports

    Article Title: Benchmarking of the Oxford Nanopore MinION sequencing for quantitative and qualitative assessment of cDNA populations

    doi: 10.1038/srep31602

    Figure Lengend Snippet: Effect of the GC content and ERCC length on the estimation of the ERCC cDNA abundance with the ONT MinION platform. The figures present deviations of the ERCC expression level estimates with the ONT MinION platform from the Ambion RNA molecular counts ( A,C ) or from the Illumina HiSeq 2500/MiSeq estimated cDNA abundance ( B,D ) as a function of the GC content ( A,B ) and the ERCC length ( C,D ). We plot the log2 ratio of observed (ONT MinION) to expected (Ambion, Illumina) read counts for the ERCC spike-ins (y-axis, log) for each of the samples relative to their length or GC content (x-axis). Due to the variable sequencing depth from each ERCC MinION experiment, each point is the average value from different MinION flow cell runs if at least 5 reads have been detected for this point in the corresponding MinION runs. The points are colored differently based on the number of flow cell runs in which they were detected (red, green, cyan, purple correspond to values derived from one, two, three or four flow cell runs respectively). The standard deviation is also presented for points with values from two or more MinION runs.

    Article Snippet: For sequencing on the Illumina HiSeq 2500 or MiSeq platforms, the cDNA was tagmented using the Nextera XT DNA Library Preparation Kit (Illumina Inc) as described in .

    Techniques: Expressing, Sequencing, Flow Cytometry, Derivative Assay, Standard Deviation

    Estimation of the ERCC cDNA abundance with the ONT MinION platform. We compared the ERCC cDNA abundance estimated from the ONT MinION ( A ) and the Illumina HiSeq 2500 or MiSeq platforms ( B ) against the expected number of RNA molecules as provided from the manufacturer (Ambion). The template reads from the ERCC MinION experiments number 1, 2, 3, 4 were pooled together and used for ( A ). Similarly, the corresponding Illumina data were pooled together for ( B ). The Illumina molecular counts data were derived using 5′ molecular tags at the RT step as described in material and methods. The total number of molecules presented on the x-axis corresponds to 3.5 pgs of ERCC RNA. In both axes the log10 transformation of the original count number is used.

    Journal: Scientific Reports

    Article Title: Benchmarking of the Oxford Nanopore MinION sequencing for quantitative and qualitative assessment of cDNA populations

    doi: 10.1038/srep31602

    Figure Lengend Snippet: Estimation of the ERCC cDNA abundance with the ONT MinION platform. We compared the ERCC cDNA abundance estimated from the ONT MinION ( A ) and the Illumina HiSeq 2500 or MiSeq platforms ( B ) against the expected number of RNA molecules as provided from the manufacturer (Ambion). The template reads from the ERCC MinION experiments number 1, 2, 3, 4 were pooled together and used for ( A ). Similarly, the corresponding Illumina data were pooled together for ( B ). The Illumina molecular counts data were derived using 5′ molecular tags at the RT step as described in material and methods. The total number of molecules presented on the x-axis corresponds to 3.5 pgs of ERCC RNA. In both axes the log10 transformation of the original count number is used.

    Article Snippet: For sequencing on the Illumina HiSeq 2500 or MiSeq platforms, the cDNA was tagmented using the Nextera XT DNA Library Preparation Kit (Illumina Inc) as described in .

    Techniques: Derivative Assay, Transformation Assay

    a Comparison of beta diversity results for the mock bacterial community and the healthy volunteer skin samples sequenced on both Roche 454 and Illumina MiSeq using Procrustes plot comparing the principal co-ordinates of Bray-Curtis distances. b Comparison of beta diversity results for the mock bacterial community and the healthy volunteer skin samples sequenced on both Roche 454 and Illumina MiSeq using Procrustes plot comparing the principal co-ordinates of Unweighted UniFrac distances. Lines connect paired sample sequences on the Roche454 ( white tip of line ) and Illumina Miseq ( red tip of line )

    Journal: BMC Microbiology

    Article Title: Optimisation of methods for bacterial skin microbiome investigation: primer selection and comparison of the 454 versus MiSeq platform

    doi: 10.1186/s12866-017-0927-4

    Figure Lengend Snippet: a Comparison of beta diversity results for the mock bacterial community and the healthy volunteer skin samples sequenced on both Roche 454 and Illumina MiSeq using Procrustes plot comparing the principal co-ordinates of Bray-Curtis distances. b Comparison of beta diversity results for the mock bacterial community and the healthy volunteer skin samples sequenced on both Roche 454 and Illumina MiSeq using Procrustes plot comparing the principal co-ordinates of Unweighted UniFrac distances. Lines connect paired sample sequences on the Roche454 ( white tip of line ) and Illumina Miseq ( red tip of line )

    Article Snippet: The mock bacterial community had an SW diversity index (n = 4 for each platform) of 1.85 and 2 for the Roche 454 Junior and the Illumina MiSeq platforms, respectively, when calculated based on the genus level distribution (Fig.

    Techniques:

    Heatmap of all 97% operational taxonomic units ( OTU s) that had at least 5% relative abundance in one sample of the diet experiment data set generated on the Illumina MiSeq platform. Colours in each cell reveal relative abundances of the identified OTU s within each sample (columns), and cells are not outlined if the OTU s were absent. Sample columns are ordered by species pair, nest ID and treatment group, while OTU s are listed in rows and ordered based on the bacterial phylogenetic tree given towards the left. OTU classifications and the total number of assigned reads per OTU across the 36 samples are given towards the right and are colour‐coded by bacterial order, as shown in the bacterial phylogeny. Asterisks show the OTU s that were consistently shared by hosts and parasites across all treatment groups in a least one nest. Arrow direction highlights the OTU s that substantially increased or decreased between the start of the experiment and the end of the sucrose treatment as shown by the similarity percentages test ( SIMPER ; for complete results see Table S9, Supporting information).

    Journal: Molecular Ecology

    Article Title: Bacterial symbiont sharing in Megalomyrmex social parasites and their fungus‐growing ant hosts

    doi: 10.1111/mec.13216

    Figure Lengend Snippet: Heatmap of all 97% operational taxonomic units ( OTU s) that had at least 5% relative abundance in one sample of the diet experiment data set generated on the Illumina MiSeq platform. Colours in each cell reveal relative abundances of the identified OTU s within each sample (columns), and cells are not outlined if the OTU s were absent. Sample columns are ordered by species pair, nest ID and treatment group, while OTU s are listed in rows and ordered based on the bacterial phylogenetic tree given towards the left. OTU classifications and the total number of assigned reads per OTU across the 36 samples are given towards the right and are colour‐coded by bacterial order, as shown in the bacterial phylogeny. Asterisks show the OTU s that were consistently shared by hosts and parasites across all treatment groups in a least one nest. Arrow direction highlights the OTU s that substantially increased or decreased between the start of the experiment and the end of the sucrose treatment as shown by the similarity percentages test ( SIMPER ; for complete results see Table S9, Supporting information).

    Article Snippet: H.L. and S.J.S. performed sequencing runs on the Roche Life Sciences 454 GS FLX Titanium and MiSeq Illumina platforms.

    Techniques: Generated