Whole-exome sequencing with at least 97% coverage at 20x was performed using the Illumina NovaSeq6000 System (Illumina, San Diego, CA, USA). Library preparation was performed using the Illumina Exome Panel (Illumina) according to the manufacturer's protocol. Library enrichment was tested by qPCR, and the size distribution and concentration were determined using Agilent Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA). The NovaSeq6000 System (Illumina) was used for DNA sequencing through 150 bp paired-end reads. Variant calling was performed according to the GATK4 (O’Connor and Auwera 2020 ) best practice guidelines, using BWA (Li and Durbin 2010 (link)) for mapping and ANNOVAR (Wang et al. 2010 (link)) for annotating.
Novaseq 6000 system
Manufactured by Illumina
Sourced in United States, China, United Kingdom, Germany, Italy, Sweden, Japan, Switzerland
The NovaSeq 6000 system is a high-throughput next-generation sequencing (NGS) platform designed for large-scale genomic research. It is capable of generating high-quality sequencing data across a wide range of applications, including whole-genome sequencing, exome sequencing, and targeted gene panels.
Automatically generated - may contain errors
Lab products found in correlation
Agilent 2100 bioanalyzer, by Agilent Technologies (58 mentions)
Trizol reagent, by Thermo Fisher Scientific (58 mentions)
2100 bioanalyzer, by Agilent Technologies (42 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (36 mentions)
Rneasy mini kit, by Qiagen (30 mentions)
Qubit 2.0 fluorometer, by Thermo Fisher Scientific (26 mentions)
Qubit dsdna hs assay kit, by Thermo Fisher Scientific (20 mentions)
Bioanalyzer 2100 system, by Agilent Technologies (18 mentions)
Nanodrop 2000 spectrophotometer, by Thermo Fisher Scientific (18 mentions)
Trizol, by Thermo Fisher Scientific (18 mentions)
Qubit 3.0 fluorometer, by Thermo Fisher Scientific (17 mentions)
Bioanalyzer, by Agilent Technologies (16 mentions)
Chromium controller, by 10x Genomics (16 mentions)
Truseq stranded mrna lt sample prep kit, by Illumina (14 mentions)
Agilent bioanalyzer 2100 system, by Agilent Technologies (14 mentions)
Qubit fluorometer, by Thermo Fisher Scientific (14 mentions)
Nebnext poly a mrna magnetic isolation module, by New England Biolabs (12 mentions)
Rq1 dnase, by Promega (12 mentions)
Truseq stranded mrna library prep kit, by Illumina (11 mentions)
Ampure xp bead, by Beckman Coulter (11 mentions)
1 021 protocols using novaseq 6000 system
1
Whole-Exome Sequencing with NovaSeq6000
2
Streamlined RNA-seq Library Preparation from Mouse Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
RNA-seq libraries from PRC1ctrl and PRC1cKO Aundiff were prepared as follows using the Smart-Seq2 method. ∼7000 Aundiff cells were isolated from 2-month-old mice and pooled as one replicate. Two independent biological replicates were used for RNA-seq library generation. Total RNA was extracted using the RNeasy Plus Micro Kit (QIAGEN, Cat #74034) according to the manufacturer's instructions. Libraries were constructed using NEBNext® Single Cell/Low Input RNA Library Prep Kit for Illumina® (NEB, E6420S) according to the manufacturer's instructions. Prepared RNA-seq libraries were sequenced on the Novaseq 6000 system (Illumina) with 100-bp paired-end reads.
Stranded RNA-seq libraries were prepared for (i) WT Aundiff and WT Adiff; (ii) PRC1ctrl Aundiff and PRC1cKO Aundiff after Win18,446 treatment. ∼100 000–150 000 cells isolated from adult mice were pooled as one replicate, and two independent biological replicates were used for RNA-seq library generation. Total RNA was extracted using the RNeasy Plus Mini Kit (QIAGEN, Cat #74134) according to the manufacturer's instructions. Library construction was performed by the CCHMC DNA Sequencing and Genotyping Core (Cincinnati, Ohio, USA) using the Illumina TruSeq stranded mRNA kit after polyA enrichment. Prepared RNA-seq libraries were sequenced on the Novaseq 6000 system (Illumina) with 100-bp paired-end reads.
Stranded RNA-seq libraries were prepared for (i) WT Aundiff and WT Adiff; (ii) PRC1ctrl Aundiff and PRC1cKO Aundiff after Win18,446 treatment. ∼100 000–150 000 cells isolated from adult mice were pooled as one replicate, and two independent biological replicates were used for RNA-seq library generation. Total RNA was extracted using the RNeasy Plus Mini Kit (QIAGEN, Cat #74134) according to the manufacturer's instructions. Library construction was performed by the CCHMC DNA Sequencing and Genotyping Core (Cincinnati, Ohio, USA) using the Illumina TruSeq stranded mRNA kit after polyA enrichment. Prepared RNA-seq libraries were sequenced on the Novaseq 6000 system (Illumina) with 100-bp paired-end reads.
3
Illumina NovaSeq6000 WES Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
WES with at least 97% coverage at 20× was performed using the Illumina NovaSeq6000 System (Illumina, San Diego, CA, USA).
Sample preparation was performed following the Nextera Flex for Enrichment manufacturer protocol. The workflow uses a bead-based transposome complex to tagment genomic DNA, which is a process that fragments DNA and then tags the DNA with adapter sequences in one step. After it is saturated with input DNA, the bead-based transposome complex fragments a set number of DNA molecules. This fragmentation provides flexibility to use a wide DNA input range to generate normalized libraries of consistent tight fragment size distribution. Following tagmentation, a limited-cycle PCR adds adapter sequences to the ends of a DNA fragment. A subsequent target enrichment workflow is then applied. Following pooling, the double stranded DNA libraries are denatured and biotinylated Illumina CEX Panel probes are hybridized to the denatured library fragments. After hybridization, Streptavidin Magnetic Beads then capture the targeted library fragments within the regions of interest. The captured and indexed libraries are eluted from beads and further amplified before sequencing. The WES analysis was performed on the Illumina NovaSeq6000 System (Illumina San Diego, CA, USA) according to the NovaSeq6000 System Guide.
Sample preparation was performed following the Nextera Flex for Enrichment manufacturer protocol. The workflow uses a bead-based transposome complex to tagment genomic DNA, which is a process that fragments DNA and then tags the DNA with adapter sequences in one step. After it is saturated with input DNA, the bead-based transposome complex fragments a set number of DNA molecules. This fragmentation provides flexibility to use a wide DNA input range to generate normalized libraries of consistent tight fragment size distribution. Following tagmentation, a limited-cycle PCR adds adapter sequences to the ends of a DNA fragment. A subsequent target enrichment workflow is then applied. Following pooling, the double stranded DNA libraries are denatured and biotinylated Illumina CEX Panel probes are hybridized to the denatured library fragments. After hybridization, Streptavidin Magnetic Beads then capture the targeted library fragments within the regions of interest. The captured and indexed libraries are eluted from beads and further amplified before sequencing. The WES analysis was performed on the Illumina NovaSeq6000 System (Illumina San Diego, CA, USA) according to the NovaSeq6000 System Guide.
4
Genetic Factors in COVID-19 Severity
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Within the GEN-COVID Multicenter Study, biospecimens from more than 3,000 SARS-CoV-2-positive individuals were collected in the GEN-COVID Biobank (GCB) and used for identifying multiorgan involvement in COVID-19, defining genetic parameters for infection susceptibility within the population and mapping genetically COVID-19 severity and clinical complexity among patients. In particular, within the GEN-COVID Multicenter Study, about 3,000 patients were sequenced by whole-exome sequencing (WES) and partly (about 2,000) already included in the model described in Fallerini et al. (2022) . WES with at least 97% coverage at 20x was performed using the Illumina NovaSeq 6000 System (Illumina, San Diego, CA, United States). Library preparation was performed using the Illumina Exome Panel (Illumina) according to the manufacturer's protocol. Library enrichment was tested by qPCR, and the size distribution and concentration were determined using the Agilent Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, United States). The NovaSeq 6000 System (Illumina) was used for DNA sequencing through 150 bp paired-end reads. Variant calling was performed according to the GATK4 (O'Connor and Auwera 2020) best practice guidelines, using BWA (Li and Durbin 2010) for mapping and ANNOVAR (Wang et al., 2010) for annotating.
5
Transcriptomic analysis of peripheral T-cell lymphoma
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was extracted from frozen tumor samples of 186 PTCL patients using RNeasy Mini Kit (Qiagen, Hilden, Germany). RNA size, concentration, and integrity were verified using Agilent 2100 System (Agilent). RNA libraries were constructed by using VAHTS total RNA-seq (HMR) library prep kit according to the manufacturer’s instructions, and sequenced on Illumina NovaSeq 6000 System. Paired-end reads were harvested from Illumina NovaSeq 6000 System, and the quality were controlled by Q30. After 3′ adaptor-trimming and low-quality reads removing by Trimmomatic software (v0.36), the high-quality trimmed reads were aligned to the reference genome (UCSC hg19) guided by the Ensembl GFF gene annotation file with STAR software (v2.5.3a).
R package limma (v3⋅38⋅3) was used to normalize raw reads and obtain differentially expressed genes. GSEA was performed using the BROAD Institute GSEA software (http://www.broad.mit.edu/gsea/ ) referring to Kyoto Encyclopedia of Genes and Genomes (KEGG) and Reactome databases.48 (link),67 (link)
R package limma (v3⋅38⋅3) was used to normalize raw reads and obtain differentially expressed genes. GSEA was performed using the BROAD Institute GSEA software (
6
RNA-seq analysis of mutant Jurkat cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
TET2mutRHOAmut, TET2mutRHOAwt, and KMT2Cmut Jurkat cells were treated with different agents. RNA size, concentration, and integrity were verified using Agilent 4200 TapeStation System (Agilent). RNA libraries were constructed by using KAPA RNA HyperPrep Kit with RiboErase (HMR, KAPA) according to the manufacturer’s instructions. Libraries were sequenced on Illumina NovaSeq 6000 System. Paired-end reads were harvested from Illumina NovaSeq 6000 System, and were quality controlled by Q30. After 3′adaptor-trimming and low-quality reads removing by cutadapt software (v1.9), the high-quality trimmed reads were aligned to the reference genome (UCSC hg19) guided by the Ensembl GFF gene annotation file with hisat2 software (v2.0.4). The GFOLD (generalized fold change) algorithm was used to find biologically meaningful rankings of differentially expressed genes from RNA-seq data of cell lines.65 (link) DEGs were then analyzed by the Database for Annotation, Visualization and Integrated Discovery (DAVID) and were enriched in KEGG pathways.
7
Single-Cell Analysis of Osteoporosis and Atherosclerosis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The single-cell data in our study were obtained from the Gene Expression Omnibus (GEO) database, and the data sources are summarized in Table 1 https://ftp.ncbi.nlm.nih.gov/geo/samples/GSM4423nnn/GSM4423510/ ) (12 (link), 13 ), which was derived from a bone marrow biopsy from a 67-year-old postmenopausal osteoporotic patient. CD271+ bone marrow-derived mononuclear cells (BM-MNCs) were extracted from the tissue and sequenced on an Illumina NovaSeq 6000 system. Carotid atherosclerosis scRNA transcriptome sequencing data were obtained from GSM4705591 (https://ftp.ncbi.nlm.nih.gov/geo/samples/GSM4705nnn/GSM4705591/ ) (11 (link), 14 ), which was derived from the carotid artery obtained from an endarterectomy of a 76-year-old postmenopausal patient with carotid atherosclerosis. Single cells were digested from atherosclerotic plaques of carotid arteries and sequenced on an Illumina NovaSeq 6000 system.
8
m6A-seq for RNA Methylation Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
m6A-seq libraries were prepared with the use of an EpiNext CUT&RUN RNA m6A-Seq Kit (EpiGentek). In brief, total RNA (5 µg) extracted from mouse heart was subjected to immunoprecipitation with the m6A antibody (P-9016, EpiGentek, 1:100 dilution) and cleaved on beads. The beads were then washed, RNA was purified from the beads and subjected to RT, and the resulting cDNA was amplified by PCR. The libraries were sequenced with a NovaSeq 6000 system (Illumina).
9
Transcriptional Analysis of Tumor Biopsies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Transcriptional analysis of the patient tumor biopsies has been described elsewhere (44 (link)). Briefly, patient tumor biopsies were either fresh-frozen and stored in liquid nitrogen or immediately treated with RNAlater (Thermo Fisher Scientific) and stored at –80°C. Each RNAlater-treated sample was homogenized with the QIAGEN TissueRuptor II, and RNA was isolated with the AllPrep DNA/RNA Mini Kit (QIAGEN). The NEBNext Ultra II RNA Library Prep Kit for Illumina was used to prepare sequencing libraries. Paired-end sequencing (PE150) was performed on the NovaSeq 6000 system (Illumina). Sequences were mapped and quantified to the decoy-aware, concatenated transcriptome of GRCh38.p13 (Ensembl, version 102) and MCPyV (R17b) using Salmon (78 (link)). Gene-level counts were generated via TxImport (79 (link)), and normalized counts were generated via DESeq2 (80 (link)). Samples were considered virus positive if the number of normalized MCPyV LT counts was greater than 100 and the number of normalized ST counts was greater than 10.
10
Metagenomic Sequencing of Gut Microbiomes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Metagenomic sequence data were collected by not only downloading previously deposited data of pig, mouse, and dog gut microbiomes from the NCBI SRA database (Xiao et al., 2015 (link); Rosshart et al., 2017 (link); Coelho et al., 2018 (link); Munk et al., 2018 (link)), but also by directly sequencing 20 pig samples (Supplementary Table 6 ).
All rectal grab fecal samples were collected aseptically from individual pigs on the same day at 75 days (30 kg) and 150 days (90 kg) of age. Total DNA from fecal samples was isolated using the FastDNA Spin Kit for Soil (MP Biomedicals), and its quality was checked using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific). DNA libraries for whole-genome shotgun sequencing were prepared according to the Illumina TruSeq Nano protocol, and 2 × 100 paired-end sequencing was performed using an Illumina NovaSeq 6000 system.
All rectal grab fecal samples were collected aseptically from individual pigs on the same day at 75 days (30 kg) and 150 days (90 kg) of age. Total DNA from fecal samples was isolated using the FastDNA Spin Kit for Soil (MP Biomedicals), and its quality was checked using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific). DNA libraries for whole-genome shotgun sequencing were prepared according to the Illumina TruSeq Nano protocol, and 2 × 100 paired-end sequencing was performed using an Illumina NovaSeq 6000 system.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!