Total DNA from E. coli ST131 strains FV9873, E35BA, E2022 and E61BA was extracted with QIAmp DNA Mini Kit (Qiagen). DNA concentration was measured with Nanodrop 2000 (Thermo Scientific) and Qubit 2.0 Fluorometer (Life Technologies). 1.0 µg DNA was sonicated (20 cycles of 30 s at 4°C, low intensity) with Bioruptor Next Generation (Diagenode). Sample quality was checked in a Bioanalyzer 2100 (Agilent Technologies). DNA samples were preconditioned for sequencing by using the TruSeq DNA Sample Preparation Kit (Illumina) and quantified with Step One Plus Real-Time PCR System (Applied Biosystems). Flow-cells were prepared with TruSeq PE Cluster Kit v5-CS-GA (Illumina). Sequencing was carried out using a standard 2×71 base protocol (300-400 bp insert size) in a Genome Analyzer IIx (Illumina, San Diego, CA) at the sequencing facility of the University of Cantabria. The main statistics of the eight sequence datasets analyzed are shown in Table 4 .
Truseq pe cluster kit v5 cs ga
Manufactured by Illumina
The TruSeq PE Cluster Kit v5-CS-GA is a laboratory equipment product designed for use with Illumina sequencing platforms. The core function of this kit is to generate clusters of DNA fragments on a flow cell, which is a critical step in the Illumina sequencing process.
Automatically generated - may contain errors
Lab products found in correlation
Steponeplus real time pcr system, by Thermo Fisher Scientific (1 mentions)
Genome analyzer iix, by Illumina (1 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (1 mentions)
2100 bioanalyzer, by Agilent Technologies (1 mentions)
Truseq dna sample preparation kit, by Illumina (1 mentions)
Qubit 2.0 fluorometer, by Thermo Fisher Scientific (1 mentions)
Qiamp dna mini kit, by Qiagen (1 mentions)
Genome analyzer 2 system, by Illumina (1 mentions)
Mrna seq sample preparation kit, by Illumina (1 mentions)
Truseq sbs kit v5 ga, by Illumina (1 mentions)
Rnaiso reagent, by Takara Bio (1 mentions)
Gaiix instrument, by Illumina (1 mentions)
Agilent 2100 bioanaylzer, by Agilent Technologies (1 mentions)
3 protocols using truseq pe cluster kit v5 cs ga
1
Illumina Sequencing of E. coli ST131 Strains
2
RNA-seq Library Preparation and Sequencing
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Equal amounts of total RNA (2 μg) from three individual birds were pooled within each treatment as described previously with minor modification [32 ]. The mRNA-seq libraries were constructed from 6 μg pooled RNA using the Illumina mRNA-seq sample preparation kit (Cat #: RS-930-1001, Illumina Inc., San Diego, CA, USA). For each treatment and each organ, two sequencing libraries were constructed from the pooled RNA samples. The rationale for pooling RNA samples from individual samples is that RNA pooling is cost-effective and can basically obtain genome-wide information about potentially functionally relevant variations [33 (link),34 (link)]. In addition, one purpose of the RNA-seq analysis in the current study was for comprehensive screening, and some identified genes were also examined for their expression by qPCR assays. Each library was loaded into a single lane of the Illumina Genome Analyzer II system (Illumina Inc.) and subjected to paired-end sequencing performed with the corresponding kits (TruSeq PE Cluster kit v5-CS-GA, Cat #: PE-203-5001 and TruSeq SBS Kit v5-GA, Cat #: FC-104-5001, Illumina Inc.).
3
RNA Extraction and Sequencing of Mouse Heart
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The RNA was extracted using RNAiso Reagent (Takara Biotechnology Co., Ltd.) according to the manufacturer's instructions. The concentration and purity of RNA were determined by Agilent 2100 Bioanaylzer (Agilent Technologies, Inc.). All the samples had RNA integrity numbers of >8.5 and concentration >200 ng/µl. Sequencing libraries were generated using TIANSeq Fast RNA Library Kit for Illumina® (cat. no. NR102; Tiangen Biotech Co., Ltd.) following the manufacturer's protocols. After completing the library, paired-end (150 bp) sequencing of the cDNA libraries (10 nM) was performed on the GAIIx instrument (Illumina, Inc.) using the reagents provided in the TruSeq PE Cluster Kit v5-CS-GA (cat. no. PE-203-5001; Illumina, Inc.). A standard analysis protocol was performed (32 ), and sequencing data were aligned to the hg19 human genome using TopHat version 2.06(33 (link)) and Bowtie2 2.0.0(34 (link)) and mapped to Ensembl transcripts (http://www.ensembl.org ). NovelBio Bio-Pharm Technology Co., Ltd. (http://www.novelbio.com/ ) performed the library construction and sequence using LPS-treated and untreated mouse heart samples.
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