ChIP-seq libraries were prepared from a total of 10 ng DNA using TruSeq Nano DNA Sample Prep Kit (Illumina) according to the manufacturer’s instructions. The completed libraries were quantified by 2100 Bioanalyzer (Agilent, Waldbronn, Germany). The libraries were then sequenced by running 2 × 150 cycles on the Illumina HiSeq 4000 following the HiSeq 3000/4000 SBS Kit protocol (Illumina). After the sequencing platform generated the sequencing images, the image analysis and base calling stages were performed using Off-Line Basecaller software V1.8. Sequence quality was examined using the FastQC software. After passing the Solexa CHASTITY quality filter, the clean reads were aligned to the Rat genome (UCSC RN5) using BOWTIE software V2.1.0.20 (link) The MACS V1.4.2 program21 (link) was then used for peak calling of the ChIP enrichment regions relative to the control dataset generated from input samples. The peaks in samples were annotated by the nearest gene using the newest UCSC RefSeq database. We performed an integrated analysis of KEGG (Kyoto Encyclopedia of Genes and Genomes Analysis) and GO (Gene Ontology Analysis) to find target Genes related to specific pathways.
Truseq nano dna sample prep kit
Manufactured by Illumina
Sourced in United States
The TruSeq Nano DNA Sample Prep Kit is a laboratory equipment product designed for preparing DNA samples for sequencing. It provides a streamlined workflow for DNA fragmentation, end-repair, A-tailing, and adapter ligation. The kit includes all the necessary reagents and consumables required for these processes.
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Lab products found in correlation
Hiseq 4000, by Illumina (13 mentions)
Sample purification beads, by Illumina (12 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (12 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (11 mentions)
S220 sonicator, by Covaris (10 mentions)
Hiseq 3000 4000 sbs kit, by Illumina (8 mentions)
Qubit 2.0 fluorometer dsdna hs assay, by Thermo Fisher Scientific (7 mentions)
Hiseq 3000 4000 pe cluster kit, by Illumina (5 mentions)
Hiseq x hd pe cluster kit, by Illumina (5 mentions)
Hiseq system, by Illumina (5 mentions)
2100 bioanalyzer, by Agilent Technologies (4 mentions)
Miseq system, by Illumina (4 mentions)
Qubit 2.0 fluorometer, by Thermo Fisher Scientific (4 mentions)
Qubit 3.0 fluorometer, by Thermo Fisher Scientific (4 mentions)
Maxwell 16 tissue dna purification kit, by Promega (4 mentions)
Hiseq x platform, by Illumina (3 mentions)
Ez dna methylation, by Zymo Research (3 mentions)
Novaseq 6000 platform, by Illumina (3 mentions)
Dneasy blood tissue kit, by Qiagen (3 mentions)
Hiseq 4000 platform, by Illumina (3 mentions)
72 protocols using truseq nano dna sample prep kit
1
ChIP-seq Library Preparation and Analysis
2
Illumina DNA Library Preparation
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DNA samples were end‐repaired, A‐tailed, and adaptor‐ligated using TruSeq Nano DNA Sample Prep Kit (#FC‐121‐4002, Illumina), following manufacturer's instructions. Approximately200‐1500 bp fragments were size selected using AMPure XP beads. The final size of the library was confirmed by Agilent 2100 Bioanalyzer. Samples were diluted to a final concentration of 8 pmol/L and cluster generation was performed on the Illumina cBot using HiSeq 3000/4000 PE Cluster Kit (#PE‐410‐1001, Illumina), following manufacturer's instructions. Sequencing was performed on Illumina HiSeq 4000 using HiSeq 3000/4000 SBS Kit (300 cycles) (#FC‐410–1003, Illumina), according to the manufacturer's instructions.
3
Identifying Sulphite-Tolerant Genotypes
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The K-means clustering algorithm used to identify maximally divergent genotypes was implemented in R (R Development Core Team 2011 ). Sulphite tolerance was assayed by plating onto YPD with either 10, 15 or 20 mM sodium metabisulphite in triplicate and scoring the growth of colonies as full, partial or none after 2 days at 28°C. Each strain was propagated in YPD, and high molecular weight genomic DNA was extracted using the Qiagen Blood & Cell Culture DNA Kit. Libraries were constructed using the Illumina TruSeq Nano DNA Sample Prep Kit with 550 bp insert size. Sequencing was carried out at the Beijing Genomics Institute (China) on a single 150-bp paired-end lane of an Illumina HiSeq 2000.
4
ChIP-Seq Library Preparation and Analysis
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ChIP-Seq libraries were prepared from a total of 10 ng DNA using TruSeq Nano DNA Sample Prep Kit (Illumina, USA) according to the manufacturer’s instructions. The completed libraries were quantified by 2100 Bioanalyzer (Agilent, Waldbronn, Germany). The libraries were then sequenced by running 2 × 150 cycles on the Illumina HiSeq 4000 following the HiSeq 3000/4000 SBS Kit protocol (Illumina). After the sequencing platform generated the sequencing images, the stages of image analysis and base calling were performed using Off-Line Basecaller software V1.8. Sequence quality was examined using the FastQC software. After passing Solexa CHASTITY quality filter, the clean reads were aligned to Rat genome (UCSC RN5) using BOWTIE software V2.1.0.21 (link) The MACS V1.4.2 program22 (link) was then used for peak calling of the ChIP enrichment regions relative to control data set that was generated from input samples. The peaks in samples were annotated by the nearest gene using the newest UCSC RefSeq database.
5
Illumina TruSeq Nano DNA Sequencing
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Sequencing libraries for each of the isolates were prepared using the Illumina TruSeq Nano DNA Sample Prep Kit, according to the manufacturer’s recommended protocol for sequencing of genomic DNA. The target DNA fragment size selected for library construction was 550 base pairs (bp). All the samples were sequenced on an in-house Illumina MiSeq using version 3 chemistry to obtain paired-end sequence fragments of 300 × 300 bp. Base calling and image analysis was performed with the MiSeq Control Software (MCS) version 2.3.0.
6
Brachyury ChIP-seq in Lung Cancer Cells
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To explore the underlying mechanisms of brachyury in lung cancer cells, Chromatin immunoprecipitation and sequencing (ChIP-seq) using wildtype MDA-MB-231 cells was performed. The ChIP assay kit (Millipore) was used to perform the ChIP assay. The anti-Bry antibodies used in this assay were purchased from R&D Systems (Bio-Techne, Minneapolis, MN). The Qubit® Fluorometer was used to determine the purity and concentration of DNA samples. TruSeq Nano DNA Sample Prep Kit (#FC-121–4002, Illumina, San Diego, CA) was used to end repair, tail and adaptor ligate DNA samples. AMPure XP beads were used to select the fragments of ~200–1,500 bp. The samples were diluted to a final concentration of 8 pM and cluster generation was then performed on the Illumina cBot using a HiSeq 3000/4000 PE Cluster Kit (#PE-410–1001, Illumina). Last, HiSeq 3000/4000 SBS Kit (300 cycles; #FC-410–1003, Illumina) was used to perform the sequencing on an Illumina HiSeq 4000. The data were then collected and analyzed.
7
ChIP-seq Analysis of H3K27Ac in Rat Islets
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Isolated rat islets incubated with or without 5 mM SB for 24 h were crosslinked with 1% formaldehyde, lysed, and sonicated to shear the chromatin into appropriate fragments. Immunoprecipitation was performed with the H3K27Ac antibody (Millipore). TruSeq Nano DNA Sample Prep Kit (Illumina) was used for sequencing library preparation. Sequencing was performed on Illumina HiSeq 4000 using HiSeq 3000/4000 SBS Kit for 300 cycles. MACS v1.4.2 (Model-based Analysis of ChIP-seq) software was run with the mapped reads to detect the statistically significant ChIP-enriched peaks compared with the respective input group by a p-value threshold of 10−4. All regions were annotated by the gene whose TSS was nearest to the center of the peak region according to the newest UCSC RefSeq database and divided into five classes based on the distance to UCSC RefSeq genes. Coverage, reads, and peaks were visualized with Integrative Genomics Viewer (IGV).
8
Hi-C Sequencing of Neuronal Progenitor Cells
9
Genomic Sequencing of Saccharomyces cerevisiae from New Zealand Vineyards
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The sampling locations of the 104 strains were across major wine growing areas on NZ’s north and south islands (Supplementary Table S1 ), and all but one strain derived from spontaneous wine ferments or vineyard habitats; the one exception was isolated from native forest soil. Fifty-two unsequenced strains were selected to complement those genomes analyzed by Gayevskiy et al. (2016) (link) (originally reported in Goddard et al. 2010 (link); Knight and Goddard 2015 (link)). Each strain was propagated in YPD, and high molecular weight genomic DNA was extracted using the Qiagen Blood & Cell Culture DNA Kit. Libraries were constructed using the Illumina TruSeq Nano DNA Sample Prep Kit with 550 bp insert size. Sequencing was carried out at the Beijing Genomics Institute (China) on three 150-bp paired-end lanes of an Illumina HiSeq 2000.
10
Minicircle assembly from pooled kDNA samples
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In order to generate a reference quality minicircle assembly, multiple samples were pooled to ensure good representation of even minor classes (Table 1 ). Libraries were constructed from purified kDNA using the TruSeq Nano DNA Sample Prep Kit or from kDNA mixed with total DNA from a cell line lacking kDNA (serving as ‘carrier’) using the Illumina TruSeq DNA Sample Prep Kit (see Table 1 ). In both cases, DNA was fragmented to generate ∼550 bp inserts and sequenced to generate paired-end Illumina MiSeq 300-bp reads (Edinburgh Genomics). Reads were quality checked and trimmed using fastqc (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/ ) and Trimmomatic (33 (link)) (bases below Q15 were removed). Reads corresponding to the nuclear megabase-sized chromosomes were removed by alignment to the published T. brucei TREU 927 genome (34 (link)) (v5.2; obtained from http://www.genedb.org/ ) with bowtie2 (35 (link)), using default parameters.
Scripts used for processing sequencing data in this study are available athttps://github.com/nicksavill/kDNA-annotation . This includes scripts for general bioinformatics tasks as well as specific programmes. Each script includes all the information required for appropriate use.
Scripts used for processing sequencing data in this study are available at
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