cbot Illumina Inc Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 92
    Illumina Inc illumina cbot
    Setting up lane-by-lane sequencing on an <t>Illumina</t> GAIIx a) <t>cBOT</t> cluster plate set up to cluster lanes 5–8. Lanes 1–4 have been removed using a razor blade and replaced with vials containing HSB. b) The TileSelection section of an XML recipe file for 4-lane sequencing of the flowcell clustered in a. Only lanes 5–8 will be imaged. c) Disconnecting the syringe-pumps. Once the manifold housing is removed, the plunger heads can be unscrewed from the motorized crossbar. Here, pumping has been inactivated on lanes 1 and 2. d) Pause-and-pulse reagent delivery method for reads longer than 40 nt. The left lane is active and the right lane is disconnected. Orange arrows represent flow of new reagent, green arrows represent movement of previous reagent, white circle represents a small air bubble remaining in the disconnected lane. i. During the initial pumping stage, reagent delivered from the inlet (bottom) may be contaminated by fluid from disconnected lanes as small bubbles expand due to negative pressure from the pump. ii. After pumping, a 5-second pause allows a small amount of the current reagent to back-fill through the inlet manifold as pressure equalizes in the system. iii. A 20 μL pulse then finishes delivery with homogenous reagent.
    Illumina Cbot, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 92/100, based on 29474 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/illumina cbot/product/Illumina Inc
    Average 92 stars, based on 29474 article reviews
    Price from $9.99 to $1999.99
    illumina cbot - by Bioz Stars, 2020-09
    92/100 stars
      Buy from Supplier

    94
    Illumina Inc cbot cluster generation system
    Setting up lane-by-lane sequencing on an <t>Illumina</t> GAIIx a) <t>cBOT</t> cluster plate set up to cluster lanes 5–8. Lanes 1–4 have been removed using a razor blade and replaced with vials containing HSB. b) The TileSelection section of an XML recipe file for 4-lane sequencing of the flowcell clustered in a. Only lanes 5–8 will be imaged. c) Disconnecting the syringe-pumps. Once the manifold housing is removed, the plunger heads can be unscrewed from the motorized crossbar. Here, pumping has been inactivated on lanes 1 and 2. d) Pause-and-pulse reagent delivery method for reads longer than 40 nt. The left lane is active and the right lane is disconnected. Orange arrows represent flow of new reagent, green arrows represent movement of previous reagent, white circle represents a small air bubble remaining in the disconnected lane. i. During the initial pumping stage, reagent delivered from the inlet (bottom) may be contaminated by fluid from disconnected lanes as small bubbles expand due to negative pressure from the pump. ii. After pumping, a 5-second pause allows a small amount of the current reagent to back-fill through the inlet manifold as pressure equalizes in the system. iii. A 20 μL pulse then finishes delivery with homogenous reagent.
    Cbot Cluster Generation System, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 94/100, based on 3721 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cbot cluster generation system/product/Illumina Inc
    Average 94 stars, based on 3721 article reviews
    Price from $9.99 to $1999.99
    cbot cluster generation system - by Bioz Stars, 2020-09
    94/100 stars
      Buy from Supplier

    94
    Illumina Inc truseq pe cluster kit v3 cbot hs
    Setting up lane-by-lane sequencing on an <t>Illumina</t> GAIIx a) <t>cBOT</t> cluster plate set up to cluster lanes 5–8. Lanes 1–4 have been removed using a razor blade and replaced with vials containing HSB. b) The TileSelection section of an XML recipe file for 4-lane sequencing of the flowcell clustered in a. Only lanes 5–8 will be imaged. c) Disconnecting the syringe-pumps. Once the manifold housing is removed, the plunger heads can be unscrewed from the motorized crossbar. Here, pumping has been inactivated on lanes 1 and 2. d) Pause-and-pulse reagent delivery method for reads longer than 40 nt. The left lane is active and the right lane is disconnected. Orange arrows represent flow of new reagent, green arrows represent movement of previous reagent, white circle represents a small air bubble remaining in the disconnected lane. i. During the initial pumping stage, reagent delivered from the inlet (bottom) may be contaminated by fluid from disconnected lanes as small bubbles expand due to negative pressure from the pump. ii. After pumping, a 5-second pause allows a small amount of the current reagent to back-fill through the inlet manifold as pressure equalizes in the system. iii. A 20 μL pulse then finishes delivery with homogenous reagent.
    Truseq Pe Cluster Kit V3 Cbot Hs, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 94/100, based on 2915 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/truseq pe cluster kit v3 cbot hs/product/Illumina Inc
    Average 94 stars, based on 2915 article reviews
    Price from $9.99 to $1999.99
    truseq pe cluster kit v3 cbot hs - by Bioz Stars, 2020-09
    94/100 stars
      Buy from Supplier

    95
    Illumina Inc cdna libraries
    Flowchart of the pipeline for the A. donax leaf transcriptome sequencing, de novo assembly, annotation, and analysis. The pipeline performs multiple operations from sampling and preparation to sequencing and de novo assembly to functional annotation and analysis. First, mRNA extraction from the first fully expanded leaf (5th from the top ) was carried out followed by <t>cDNA</t> preparation and library construction ( gray ). Sequencing was performed using an <t>Illumina</t> HiSeq platform. The sequenced reads were then subjected to quality control and filtering, and identical sequences were removed ( blue ). Next, de novo assemblies of transcripts were generated using a two-step approach: first, multi- k - mer (Trans-ABySS and rnaSPAdes) and single- k - mer (Trinity) methods were used to generate the pre-assemblies ( pink ); second, pre-assemblies were then concatenated and redundant transcripts were removed using CD-HIT and the EvidentialGene tr2aadcs pipeline ( purple ). The quality of the de novo assembled leaf transcriptome was then assessed ( green ). Finally, the non-redundant (NR) transcript dataset was functionally annotated by homology and gene ontology (GO), and metabolic pathways were analyzed ( mint blue ). Simple sequence repeats (SSRs) and polymorphic SSRs (PolySSRs) were also identified ( orange )
    Cdna Libraries, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 95/100, based on 30682 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cdna libraries/product/Illumina Inc
    Average 95 stars, based on 30682 article reviews
    Price from $9.99 to $1999.99
    cdna libraries - by Bioz Stars, 2020-09
    95/100 stars
      Buy from Supplier

    90
    Illumina Inc truseq pe cluster kit v4 cbot hs
    Flowchart of the pipeline for the A. donax leaf transcriptome sequencing, de novo assembly, annotation, and analysis. The pipeline performs multiple operations from sampling and preparation to sequencing and de novo assembly to functional annotation and analysis. First, mRNA extraction from the first fully expanded leaf (5th from the top ) was carried out followed by <t>cDNA</t> preparation and library construction ( gray ). Sequencing was performed using an <t>Illumina</t> HiSeq platform. The sequenced reads were then subjected to quality control and filtering, and identical sequences were removed ( blue ). Next, de novo assemblies of transcripts were generated using a two-step approach: first, multi- k - mer (Trans-ABySS and rnaSPAdes) and single- k - mer (Trinity) methods were used to generate the pre-assemblies ( pink ); second, pre-assemblies were then concatenated and redundant transcripts were removed using CD-HIT and the EvidentialGene tr2aadcs pipeline ( purple ). The quality of the de novo assembled leaf transcriptome was then assessed ( green ). Finally, the non-redundant (NR) transcript dataset was functionally annotated by homology and gene ontology (GO), and metabolic pathways were analyzed ( mint blue ). Simple sequence repeats (SSRs) and polymorphic SSRs (PolySSRs) were also identified ( orange )
    Truseq Pe Cluster Kit V4 Cbot Hs, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 90/100, based on 334 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/truseq pe cluster kit v4 cbot hs/product/Illumina Inc
    Average 90 stars, based on 334 article reviews
    Price from $9.99 to $1999.99
    truseq pe cluster kit v4 cbot hs - by Bioz Stars, 2020-09
    90/100 stars
      Buy from Supplier

    94
    Illumina Inc genome analyzer iix
    Flowchart of the pipeline for the A. donax leaf transcriptome sequencing, de novo assembly, annotation, and analysis. The pipeline performs multiple operations from sampling and preparation to sequencing and de novo assembly to functional annotation and analysis. First, mRNA extraction from the first fully expanded leaf (5th from the top ) was carried out followed by <t>cDNA</t> preparation and library construction ( gray ). Sequencing was performed using an <t>Illumina</t> HiSeq platform. The sequenced reads were then subjected to quality control and filtering, and identical sequences were removed ( blue ). Next, de novo assemblies of transcripts were generated using a two-step approach: first, multi- k - mer (Trans-ABySS and rnaSPAdes) and single- k - mer (Trinity) methods were used to generate the pre-assemblies ( pink ); second, pre-assemblies were then concatenated and redundant transcripts were removed using CD-HIT and the EvidentialGene tr2aadcs pipeline ( purple ). The quality of the de novo assembled leaf transcriptome was then assessed ( green ). Finally, the non-redundant (NR) transcript dataset was functionally annotated by homology and gene ontology (GO), and metabolic pathways were analyzed ( mint blue ). Simple sequence repeats (SSRs) and polymorphic SSRs (PolySSRs) were also identified ( orange )
    Genome Analyzer Iix, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 94/100, based on 12049 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/genome analyzer iix/product/Illumina Inc
    Average 94 stars, based on 12049 article reviews
    Price from $9.99 to $1999.99
    genome analyzer iix - by Bioz Stars, 2020-09
    94/100 stars
      Buy from Supplier

    95
    Illumina Inc rna seq libraries
    Flowchart of the pipeline for the A. donax leaf transcriptome sequencing, de novo assembly, annotation, and analysis. The pipeline performs multiple operations from sampling and preparation to sequencing and de novo assembly to functional annotation and analysis. First, mRNA extraction from the first fully expanded leaf (5th from the top ) was carried out followed by <t>cDNA</t> preparation and library construction ( gray ). Sequencing was performed using an <t>Illumina</t> HiSeq platform. The sequenced reads were then subjected to quality control and filtering, and identical sequences were removed ( blue ). Next, de novo assemblies of transcripts were generated using a two-step approach: first, multi- k - mer (Trans-ABySS and rnaSPAdes) and single- k - mer (Trinity) methods were used to generate the pre-assemblies ( pink ); second, pre-assemblies were then concatenated and redundant transcripts were removed using CD-HIT and the EvidentialGene tr2aadcs pipeline ( purple ). The quality of the de novo assembled leaf transcriptome was then assessed ( green ). Finally, the non-redundant (NR) transcript dataset was functionally annotated by homology and gene ontology (GO), and metabolic pathways were analyzed ( mint blue ). Simple sequence repeats (SSRs) and polymorphic SSRs (PolySSRs) were also identified ( orange )
    Rna Seq Libraries, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 95/100, based on 14521 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rna seq libraries/product/Illumina Inc
    Average 95 stars, based on 14521 article reviews
    Price from $9.99 to $1999.99
    rna seq libraries - by Bioz Stars, 2020-09
    95/100 stars
      Buy from Supplier

    Image Search Results


    Setting up lane-by-lane sequencing on an Illumina GAIIx a) cBOT cluster plate set up to cluster lanes 5–8. Lanes 1–4 have been removed using a razor blade and replaced with vials containing HSB. b) The TileSelection section of an XML recipe file for 4-lane sequencing of the flowcell clustered in a. Only lanes 5–8 will be imaged. c) Disconnecting the syringe-pumps. Once the manifold housing is removed, the plunger heads can be unscrewed from the motorized crossbar. Here, pumping has been inactivated on lanes 1 and 2. d) Pause-and-pulse reagent delivery method for reads longer than 40 nt. The left lane is active and the right lane is disconnected. Orange arrows represent flow of new reagent, green arrows represent movement of previous reagent, white circle represents a small air bubble remaining in the disconnected lane. i. During the initial pumping stage, reagent delivered from the inlet (bottom) may be contaminated by fluid from disconnected lanes as small bubbles expand due to negative pressure from the pump. ii. After pumping, a 5-second pause allows a small amount of the current reagent to back-fill through the inlet manifold as pressure equalizes in the system. iii. A 20 μL pulse then finishes delivery with homogenous reagent.

    Journal: BioTechniques

    Article Title: Lane-by-lane sequencing using Illumina’s Genome Analyzer II

    doi: 10.2144/000114032

    Figure Lengend Snippet: Setting up lane-by-lane sequencing on an Illumina GAIIx a) cBOT cluster plate set up to cluster lanes 5–8. Lanes 1–4 have been removed using a razor blade and replaced with vials containing HSB. b) The TileSelection section of an XML recipe file for 4-lane sequencing of the flowcell clustered in a. Only lanes 5–8 will be imaged. c) Disconnecting the syringe-pumps. Once the manifold housing is removed, the plunger heads can be unscrewed from the motorized crossbar. Here, pumping has been inactivated on lanes 1 and 2. d) Pause-and-pulse reagent delivery method for reads longer than 40 nt. The left lane is active and the right lane is disconnected. Orange arrows represent flow of new reagent, green arrows represent movement of previous reagent, white circle represents a small air bubble remaining in the disconnected lane. i. During the initial pumping stage, reagent delivered from the inlet (bottom) may be contaminated by fluid from disconnected lanes as small bubbles expand due to negative pressure from the pump. ii. After pumping, a 5-second pause allows a small amount of the current reagent to back-fill through the inlet manifold as pressure equalizes in the system. iii. A 20 μL pulse then finishes delivery with homogenous reagent.

    Article Snippet: Flowcells can be clustered on any desired number of lanes using the Illumina cBot, sequenced, stored, and then clustered again on remaining lanes without appreciable reduction in intensity, cluster density or accuracy of the run.

    Techniques: Sequencing, Flow Cytometry

    The numbers of DEGs are comparable in human dCD8 versus pCD8 T cells. (A) Fluorescence-activated cell sorting was performed to purify the CD8 + T cells, based on the phenotype of CD3 + CD8 + CD4 − , in paired decidual and peripheral blood samples from three healthy women at the first trimester of normal pregnancy (mean gestational day 50, range 44–58). (B) Summary of mRNA sequencing data of the isolated human pCD8 and autologous dCD8 T cells. 2 × 125-bp paired-end cDNA fragments were acquired after sequencing the polyadenylated [poly(A)+] enriched mRNAs on an Illumina Hiseq 2500 platform, and the sequenced reads was aligned to human reference genome (hg19 version) by STAR software. (C) The number and proportion of DEGs that were upregulated or downregulated in human dCD8 versus pCD8 T cells ( P

    Journal: Frontiers in Immunology

    Article Title: Deep Surveying of the Transcriptional and Alternative Splicing Signatures for Decidual CD8+ T Cells at the First Trimester of Human Healthy Pregnancy

    doi: 10.3389/fimmu.2018.00937

    Figure Lengend Snippet: The numbers of DEGs are comparable in human dCD8 versus pCD8 T cells. (A) Fluorescence-activated cell sorting was performed to purify the CD8 + T cells, based on the phenotype of CD3 + CD8 + CD4 − , in paired decidual and peripheral blood samples from three healthy women at the first trimester of normal pregnancy (mean gestational day 50, range 44–58). (B) Summary of mRNA sequencing data of the isolated human pCD8 and autologous dCD8 T cells. 2 × 125-bp paired-end cDNA fragments were acquired after sequencing the polyadenylated [poly(A)+] enriched mRNAs on an Illumina Hiseq 2500 platform, and the sequenced reads was aligned to human reference genome (hg19 version) by STAR software. (C) The number and proportion of DEGs that were upregulated or downregulated in human dCD8 versus pCD8 T cells ( P

    Article Snippet: After cluster generation, the prepared libraries were sequenced on an Illumina Hiseq 2500 platform and 125 bp paired-end reads were produced.

    Techniques: Fluorescence, FACS, Sequencing, Isolation, Software

    Representative terpenoid biosynthesis pathway with cognate heat maps for transcript levels of genes from transcriptome data with substrates and products, colored arrows connect substrates to their corresponding products. Green/red color-coded heat maps represent relative transcript levels of different terpenoid genes determined by Illumina HiSeq 2000 sequencing; red, upregulated; green, downregulated. Transcript levels data represent by FPKM: Fragments per Kilobase of transcripts per Million mapped fragments. MeV: MultiExperiment Viewer software was used to depict transcript levels. DXS: 1-deoxy-D-xylulose-5-phosphate synthase, DXR:1-deoxy-D-xylulose-5-phosphate reductoisomerase, MCT: 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase, ISPF: 2-C-methyl-D-erythritol 2,4-cyclodiphos-phate synthase, HDS:(E)-4-hydroxy-3-methylbut-2-enyl-diphosphate synthase, HDR: 4-hydroxy-3-methylbut-2-enyl diphosphate reductases, IDI: isopentenyl-diphosphate delta isomerase, AACT: acetyl-CoA C-acetyl transferase, HMGS: hydroxyl methyl glutaryl-CoA synthase, HMGR: hydroxymethyl glutaryl-CoA reductase (NADPH), MVK: mevalonate kinase, PMK: phospho-mevalonate kinase, GPPS: geranyl pyrophosphate synthase, FPPS: farnesyl pyrophosphate synthase, GGPS: geranylgeranyl pyrophosphate synthase, type II, CINO:1,8-cineole synthase, MYS: myrcene/ocimene synthase, LINA: (3S)-linalool synthase, NEOM:(+)-neomenthol dehydrogenase, SABI:(+)-sabinene synthase, TPS6:(−)-germacrene D synthase, AMS:beta-amyrin synthase, SEQ: Squalene monooxygenase, HUMS:α-humulene/β-caryophyllene synthase, GA2:gibberellin 2- -oxidase, GA20:gibberellin 20-oxidase, E-KS:ent-kaurene synthase, MAS:momilactone-A synthase, GA3:gibberellin 3-beta-dioxygenase, E-KIA: ent-isokaurene C2-hydroxylase, E-KIH:ent-kaurenoic acid hydroxylase, E-CDS: ent-copalyl diphosphate synthase.

    Journal: Scientific Reports

    Article Title: Transcriptome and metabolite analyses reveal the complex metabolic genes involved in volatile terpenoid biosynthesis in garden sage (Salvia officinalis)

    doi: 10.1038/s41598-017-15478-3

    Figure Lengend Snippet: Representative terpenoid biosynthesis pathway with cognate heat maps for transcript levels of genes from transcriptome data with substrates and products, colored arrows connect substrates to their corresponding products. Green/red color-coded heat maps represent relative transcript levels of different terpenoid genes determined by Illumina HiSeq 2000 sequencing; red, upregulated; green, downregulated. Transcript levels data represent by FPKM: Fragments per Kilobase of transcripts per Million mapped fragments. MeV: MultiExperiment Viewer software was used to depict transcript levels. DXS: 1-deoxy-D-xylulose-5-phosphate synthase, DXR:1-deoxy-D-xylulose-5-phosphate reductoisomerase, MCT: 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase, ISPF: 2-C-methyl-D-erythritol 2,4-cyclodiphos-phate synthase, HDS:(E)-4-hydroxy-3-methylbut-2-enyl-diphosphate synthase, HDR: 4-hydroxy-3-methylbut-2-enyl diphosphate reductases, IDI: isopentenyl-diphosphate delta isomerase, AACT: acetyl-CoA C-acetyl transferase, HMGS: hydroxyl methyl glutaryl-CoA synthase, HMGR: hydroxymethyl glutaryl-CoA reductase (NADPH), MVK: mevalonate kinase, PMK: phospho-mevalonate kinase, GPPS: geranyl pyrophosphate synthase, FPPS: farnesyl pyrophosphate synthase, GGPS: geranylgeranyl pyrophosphate synthase, type II, CINO:1,8-cineole synthase, MYS: myrcene/ocimene synthase, LINA: (3S)-linalool synthase, NEOM:(+)-neomenthol dehydrogenase, SABI:(+)-sabinene synthase, TPS6:(−)-germacrene D synthase, AMS:beta-amyrin synthase, SEQ: Squalene monooxygenase, HUMS:α-humulene/β-caryophyllene synthase, GA2:gibberellin 2- -oxidase, GA20:gibberellin 20-oxidase, E-KS:ent-kaurene synthase, MAS:momilactone-A synthase, GA3:gibberellin 3-beta-dioxygenase, E-KIA: ent-isokaurene C2-hydroxylase, E-KIH:ent-kaurenoic acid hydroxylase, E-CDS: ent-copalyl diphosphate synthase.

    Article Snippet: After cluster generation, the library preparations were sequenced on an Illumina HiSeq 2000 platform, and paired-end reads were generated.

    Techniques: Sequencing, Software, Affinity Magnetic Separation

    DNA template and RNA transcript of HiTS-RAP. (A) Schematics of the DNA template used for HiTS-RAP and the resulting halted RNA transcript. DNA template encoding the RNA of interest (green) is flanked by the Illumina flowcell adaptor 1 (gray) and T7 RNA polymerase promoter (orange) upstream, and by the Illumina sequencing primer annealing site (purple), Tus-binding Ter site (red), and Illumina flowcell adaptor 2 downstream. Illumina flowcell adaptors 1 and 2 are required for cluster generation on Illumina GA flowcell. T7 RNA polymerase promoter is required for transcription of the RNA of interest. The Illumina sequencing primer is used for sequencing the DNA template of the RNA of interest and serves as a docking site for the T7 RNA polymerase when it is halted. Tus protein binds to Ter site and halts the transcribing RNA polymerase. Direction of transcription and sequencing are indicated by orange and purple arrows, respectively. Tus-bound Ter site that is non-permissive (halting) and permissive (read-through) to RNA polymerase are indicated by solid and open red triangles, respectively. The halted RNA transcript includes a triplet G derived from the T7 promoter, followed by the RNA of interest and some of the Illumina sequencing primer. The 3’-end of RNA transcript, indicated by a dashed line, is inaccessible. (B) Construction of DNA templates for HiTS-RAP. DNA template is constructed by PCR in two steps using 2 sets of nested oligos. Forward oligos introduce T7 promoter (step 1) and Illumina flowcell adaptor (step 2), whereas reverse oligos introduce Illumina sequencing primer (step 1), and Ter site and Illumina flowcell adaptor 2 (step 2). (C) Sequence of the HiTS-RAP DNA template for GFP aptamer. GFP aptamer encoding sequence is in green, and the rest of the sequences are colored as in (A). The transcription start site is indicated by +1 and a broken arrow. (D) Sequence of the halted RNA transcript from GFP aptamer template. Sequences are colored as in (C). Uppercase indicates the region of the halted RNA transcript that is accessible, and the lowercase indicates a region that is likely to be buried in T7 RNA polymerase and thus inaccessible by other proteins.

    Journal: Nature protocols

    Article Title: Quantitative Assessment of RNA-Protein Interactions with High Throughput Sequencing - RNA Affinity Profiling (HiTS-RAP)

    doi: 10.1038/nprot.2015.074

    Figure Lengend Snippet: DNA template and RNA transcript of HiTS-RAP. (A) Schematics of the DNA template used for HiTS-RAP and the resulting halted RNA transcript. DNA template encoding the RNA of interest (green) is flanked by the Illumina flowcell adaptor 1 (gray) and T7 RNA polymerase promoter (orange) upstream, and by the Illumina sequencing primer annealing site (purple), Tus-binding Ter site (red), and Illumina flowcell adaptor 2 downstream. Illumina flowcell adaptors 1 and 2 are required for cluster generation on Illumina GA flowcell. T7 RNA polymerase promoter is required for transcription of the RNA of interest. The Illumina sequencing primer is used for sequencing the DNA template of the RNA of interest and serves as a docking site for the T7 RNA polymerase when it is halted. Tus protein binds to Ter site and halts the transcribing RNA polymerase. Direction of transcription and sequencing are indicated by orange and purple arrows, respectively. Tus-bound Ter site that is non-permissive (halting) and permissive (read-through) to RNA polymerase are indicated by solid and open red triangles, respectively. The halted RNA transcript includes a triplet G derived from the T7 promoter, followed by the RNA of interest and some of the Illumina sequencing primer. The 3’-end of RNA transcript, indicated by a dashed line, is inaccessible. (B) Construction of DNA templates for HiTS-RAP. DNA template is constructed by PCR in two steps using 2 sets of nested oligos. Forward oligos introduce T7 promoter (step 1) and Illumina flowcell adaptor (step 2), whereas reverse oligos introduce Illumina sequencing primer (step 1), and Ter site and Illumina flowcell adaptor 2 (step 2). (C) Sequence of the HiTS-RAP DNA template for GFP aptamer. GFP aptamer encoding sequence is in green, and the rest of the sequences are colored as in (A). The transcription start site is indicated by +1 and a broken arrow. (D) Sequence of the halted RNA transcript from GFP aptamer template. Sequences are colored as in (C). Uppercase indicates the region of the halted RNA transcript that is accessible, and the lowercase indicates a region that is likely to be buried in T7 RNA polymerase and thus inaccessible by other proteins.

    Article Snippet: Although all 8 lanes of the flowcell can be addressed individually on the cBot instrument during cluster generation, the Illumina GAIIx instrument processes all of them simultaneously with the same solutions at any given time.

    Techniques: Sequencing, Binding Assay, Derivative Assay, Construct, Polymerase Chain Reaction, Introduce

    Experimental design of the mixture control experiment. RNA from two lung cancer cell lines (NCI-H1975 and HCC827) were obtained after culture on three separate occasions to obtain samples for three replicates to simulate some degree of biological variability. RNA from each replicate was either kept pure or mixed in three different proportions. The second replicate of each mixture was split in two and either processed normally or heat treated (incubated at 37°C for 9 days, see Materials and Methods) to degrade the RNA and simulate variations in sample quality. Each sample was then processed using either Illumina's TruSeq RNA v2 kit (Poly-A mRNA) or Illumina's TruSeq Total Stranded RNA kit with Ribozero depletion followed by sequencing on an Illumina HiSeq 2500 to obtain 100 bp single-end reads for further analysis.

    Journal: Nucleic Acids Research

    Article Title: RNA-seq mixology: designing realistic control experiments to compare protocols and analysis methods

    doi: 10.1093/nar/gkw1063

    Figure Lengend Snippet: Experimental design of the mixture control experiment. RNA from two lung cancer cell lines (NCI-H1975 and HCC827) were obtained after culture on three separate occasions to obtain samples for three replicates to simulate some degree of biological variability. RNA from each replicate was either kept pure or mixed in three different proportions. The second replicate of each mixture was split in two and either processed normally or heat treated (incubated at 37°C for 9 days, see Materials and Methods) to degrade the RNA and simulate variations in sample quality. Each sample was then processed using either Illumina's TruSeq RNA v2 kit (Poly-A mRNA) or Illumina's TruSeq Total Stranded RNA kit with Ribozero depletion followed by sequencing on an Illumina HiSeq 2500 to obtain 100 bp single-end reads for further analysis.

    Article Snippet: Each of the two pools of libraries was sequenced as single-end 100 base pair reads over 4 lanes on an Illumina HiSeq 2500 with an Illumina HiSeq SBS kit v4.

    Techniques: Incubation, Sequencing

    Flowchart of the pipeline for the A. donax leaf transcriptome sequencing, de novo assembly, annotation, and analysis. The pipeline performs multiple operations from sampling and preparation to sequencing and de novo assembly to functional annotation and analysis. First, mRNA extraction from the first fully expanded leaf (5th from the top ) was carried out followed by cDNA preparation and library construction ( gray ). Sequencing was performed using an Illumina HiSeq platform. The sequenced reads were then subjected to quality control and filtering, and identical sequences were removed ( blue ). Next, de novo assemblies of transcripts were generated using a two-step approach: first, multi- k - mer (Trans-ABySS and rnaSPAdes) and single- k - mer (Trinity) methods were used to generate the pre-assemblies ( pink ); second, pre-assemblies were then concatenated and redundant transcripts were removed using CD-HIT and the EvidentialGene tr2aadcs pipeline ( purple ). The quality of the de novo assembled leaf transcriptome was then assessed ( green ). Finally, the non-redundant (NR) transcript dataset was functionally annotated by homology and gene ontology (GO), and metabolic pathways were analyzed ( mint blue ). Simple sequence repeats (SSRs) and polymorphic SSRs (PolySSRs) were also identified ( orange )

    Journal: Biotechnology for Biofuels

    Article Title: De novo assembly, functional annotation, and analysis of the giant reed (Arundo donax L.) leaf transcriptome provide tools for the development of a biofuel feedstock

    doi: 10.1186/s13068-017-0828-7

    Figure Lengend Snippet: Flowchart of the pipeline for the A. donax leaf transcriptome sequencing, de novo assembly, annotation, and analysis. The pipeline performs multiple operations from sampling and preparation to sequencing and de novo assembly to functional annotation and analysis. First, mRNA extraction from the first fully expanded leaf (5th from the top ) was carried out followed by cDNA preparation and library construction ( gray ). Sequencing was performed using an Illumina HiSeq platform. The sequenced reads were then subjected to quality control and filtering, and identical sequences were removed ( blue ). Next, de novo assemblies of transcripts were generated using a two-step approach: first, multi- k - mer (Trans-ABySS and rnaSPAdes) and single- k - mer (Trinity) methods were used to generate the pre-assemblies ( pink ); second, pre-assemblies were then concatenated and redundant transcripts were removed using CD-HIT and the EvidentialGene tr2aadcs pipeline ( purple ). The quality of the de novo assembled leaf transcriptome was then assessed ( green ). Finally, the non-redundant (NR) transcript dataset was functionally annotated by homology and gene ontology (GO), and metabolic pathways were analyzed ( mint blue ). Simple sequence repeats (SSRs) and polymorphic SSRs (PolySSRs) were also identified ( orange )

    Article Snippet: Finally, cDNA libraries were processed with Illumina cBot for cluster generation on the flow cell following the manufacturer’s instructions and sequenced in single-end mode using a HiSeq2500 sequencing platform (Illumina, San Diego, CA).

    Techniques: Sequencing, Sampling, Functional Assay, Generated