The high quality genomic DNA was obtained using the same method as WGBS. The DNA fragments with a growth of about 20 kb were fragmented using the g-GUBETM (520,079, Covaris) and enriched by the magnetic beads (100-317-100, Pacific Bioscience). The DNA fragments were added with stem rings to synthesize the sequences using DNA polymerase (101-731-100, Pacific Bioscience). The DNA quality was assessed and sequenced using the HiSeq X10 sequencing platform (Illumina), which produced paired 150 bp readings of the terminal.
Hiseq x10 sequencing platform
Manufactured by Illumina
Sourced in United States
The HiSeq X10 is a high-throughput DNA sequencing platform manufactured by Illumina. It is designed to deliver large-scale, whole-genome sequencing data. The HiSeq X10 can generate up to 1.8 terabases of sequencing data per run, with a read length of up to 150 base pairs.
Automatically generated - may contain errors
Lab products found in correlation
Nanodrop, by Thermo Fisher Scientific (4 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (3 mentions)
Wizard genomic dna purification kit, by Promega (2 mentions)
Nebnext ultra rna library prep kit, by Illumina (2 mentions)
Rneasy mini kit, by Qiagen (2 mentions)
G tube, by Covaris (1 mentions)
Fastrna pro red kit, by MP Biomedicals (1 mentions)
Hiseq control software olb gapipeline 1, by Illumina (1 mentions)
Qiaamp dna mini kit, by Qiagen (1 mentions)
2100 bioanalyzer, by Agilent Technologies (1 mentions)
Ez dna methylation gold kit, by Zymo Research (1 mentions)
Trizol, by Thermo Fisher Scientific (1 mentions)
Mirneasy mini kit, by Qiagen (1 mentions)
Hiseq 2500 sequencing platform, by Illumina (1 mentions)
Kapa hyper prep kit, by Roche (1 mentions)
Rnaiso plus reagent, by Takara Bio (1 mentions)
12 protocols using hiseq x10 sequencing platform
1
Genomic DNA Extraction and Sequencing
2
RNA-Seq on Illumina HiSeq X10
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RNA samples were sent to Shanghai Gminix, Biotechnology Co, Ltd. (Shanghai, China) for RNA deep sequencing on an Illumina HiSeq X10 sequencing platform with 15G PF data (Illumina, San Diego, CA). Procedures were performed as described in detail on the website of Gminix (http://www.gminix.com ). The data discussed in this publication have been deposited in NCBI sequence read archive (SRA), and are accessible through SRA Series Accession number SUB6070993.
3
Genomic Analysis of C. striatum Outbreak
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A clinical sample was obtained from hospitalized patients growing exclusively C. striatum colonies, according to the definition of an outbreak case described by Verroken et al. (14 (link)). A total of 30 C. striatum isolates were isolated from 26 patients from a hospital in Shanxi, China, between 24 March and 23 April 2021. One epidemiologically unrelated outgroup strain was collected from a hospital in Wenzhou, China, in January 2019. Genomic DNA was isolated and purified using the Wizard genomic DNA purification kit (Promega, WI, USA). Whole-genome sequencing was carried out with the Illumina HiSeq X 10 sequencing platform (150-bp paired-end reading). Trimmomatic version 0.39 (53 (link)) was used to remove adapter sequences and low-quality sequences. Assembly was performed using SPAdes v3.11.1 (54 (link)). The CheckM tool was used to assess genome completeness and contamination (55 (link)).
4
DNA Fragmentation and Sequencing Protocol
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The DNA was broken into fragments of around 20 kb using a g-TUBETM (Covaris, 520079). Following that, magnetic beads were added (Pacifi Bioscience, 100-317-100). After adding stem-loop junctions to DNA fragments, a fresh round of screening was performed. Next, DNA and primers were annealed, and DNA polymerase was used for DNA extension (Pacifi Bioscience, 101-731-100). The quality of the DNA was then assessed using an Agilent 2100 Bioanalyzer, and paired-end 150-bp reads were generated by the Hiseq X10 sequencing platform (Illumina).
5
Total RNA Extraction and Sequencing
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The total RNA of each sample was extracted by using the FastRNA Pro Red kit (MP Biomedicals, Santa Ana, CA, USA) and purified by using the RNeasy Mini Kit (Qiagen, Hilden, Germany). The RNA products obtained from the samples were quantified and qualified by using an Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA), NanoDrop (Thermo FisherScientific Inc.), and 1% agarose gel electrophoresis before next-generation sequencing of library preparations. RNA extraction, quantification, and qualification were each conducted more than three time, followed by mixing for further library preparation. Approximately 1 μg of extracted total RNA was used for library preparation, based on the manufacturer’s protocol of the NEBNext Ultra RNA Library Prep kit for Illumina. The library sequencing was conducted on the Illumina HiSeq X10 sequencing platform.
6
Yeast RNA Extraction and Sequencing
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Yeast cells were grinded in liquid nitrogen. Total RNA of each sample was extracted using RNeasy Mini Kit (Qiagen, Germany). The obtained RNA products were quantified and qualified by Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA), NanoDrop (Thermo Fisher Scientific Inc.) and 1% agarose gel electrophoresis before next generation sequencing library preparations. One microgram of total RNA passing the quality test was used for library preparation based on the manufacturer’s protocol of the NEBNext Ultra RNA Library Prep kit for Illumina.
Illumina HiSeq X10 sequencing platform was employed for the library sequencing using a 2×150 bp paired-end (PE) configuration; image analysis and base calling were conducted by the HiSeq Control Software (HCS) + OLB + GAPipeline-1.6 (Illumina) on the HiSeq instrument. The sequences were processed and analyzed by Genewiz (Suzhou, China).
Illumina HiSeq X10 sequencing platform was employed for the library sequencing using a 2×150 bp paired-end (PE) configuration; image analysis and base calling were conducted by the HiSeq Control Software (HCS) + OLB + GAPipeline-1.6 (Illumina) on the HiSeq instrument. The sequences were processed and analyzed by Genewiz (Suzhou, China).
7
Genomic DNA Extraction and Whole-Genome Sequencing
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Genomic DNA was extracted and purified using the Wizard genomic DNA purification kit (Promega, United States). Whole-genome sequencing was carried out by Illumina HiSeq X 10 sequencing platform. Removing adapter sequences and low-quality sequences was performed using Trimmomatic version 0.39. Assembly was performed with SPAdes v3.11.1.
8
Whole-genome sequencing of bacterial strains
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Genomic DNA from an overnight culture of each of the two strains was prepared using the QIAamp DNA minikit (Qiagen, Hilden, Germany) and was then subjected to whole-genome sequencing using the HiSeq X10 sequencing platform (Illumina, San Diego, CA, USA) with an approximate 250× coverage. Reads were de novo assembled into contigs using the program SPAdes v3.15.1 (44 (link)). Potential contaminations of WCHAc060012T and WCHAc060115T genomes were checked using CheckM v1.1.3 (45 (link)). Antimicrobial resistance genes were identified from genome sequences using the ABRicate program (https://github.com/tseemann/abricate ) to query the ResFinder database 4.1 (https://cge.cbs.dtu.dk/services/ResFinder/ ).
9
Genome-wide DNA Methylation Profiling
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The genomic DNA was extracted with SMRTbell Express Template Prep Kit 2.0 (100–938-900, Pacific Bioscience) and tested for quality and concentration. DNA samples were segmayed by ultrasound for bisulfite transformation, and the DNA methylation sequencing library construction Kit EZDNA methylation-Gold ™ Kit (D5005, ZYMO Research) was used to connect single strand DNA fragments for PCR amplification. The amplified products were purified and tested for their integrity using a 2,100 Bioanalyzer (Agilent). Finally, the HiSeq X10 sequencing platform (Illumina) was used for the double-terminal sequencing of the library, and the results were compared with each methylation regions.
10
RNA-seq Analysis of Silicosis Biomarkers
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Total RNA isolated from the peripheral blood leucocytes from four silicosis cases and four healthy worker controls in the first stage was extracted using TRIzol (Invitrogen) and a miRNeasy Mini Kit (QIAGEN) following the manufacturer’s instructions. Then, RNA samples were sent to Gminix, Biotechnology Co, Ltd (Shanghai, China) for RNA-seq. LncRNA, mRNA and circRNA sequencing steps were performed on an Illumina HiSeq X10 sequencing platform with 15G reads (Illumina, San Diego, California, USA), while miRNA sequencing step was performed on an Illumina HiSeq 2500 sequencing platform with 10M reads (Illumina, San Diego, California, USA).
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