Agilent sureselectxt reagent kit

Manufactured by Agilent Technologies

Sourced in United States, Switzerland

The Agilent SureSelectXT reagent kit is a laboratory equipment product designed for targeted DNA library preparation. It enables the enrichment and capture of specific genomic regions of interest, facilitating focused sequencing and analysis.

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Lab products found in correlation

Nextseq 500 device, by Illumina (4 mentions) Sureselect all exon v5 or v6, by Agilent Technologies (3 mentions) Hiseq 2500, by Illumina (2 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions) Agilent sureselect xt human all exon v6, by Agilent Technologies (1 mentions) Qubit 3 fluorometer, by Agilent Technologies (1 mentions) Novaseq 6000 s2 flow cell, by Illumina (1 mentions) Nextseq 500 high output kit, by Illumina (1 mentions) Nextseq 500 mid output kit, by Illumina (1 mentions) Qubit dsdna hs assay kit, by Thermo Fisher Scientific (1 mentions) Biorupter ultrasonicator, by Diagenode (1 mentions) M220 focused ultrasonicator, by Covaris (1 mentions) Miseq instrument, by Illumina (1 mentions)

Spelling variants of this product

SureSelectXT Reagent Kit (58 citations) SureSelectXT kit (18 citations) Agilent SureSelectXT Low Input Reagent Kit (2 citations)

10 protocols using agilent sureselectxt reagent kit

Exome Sequencing and Bioinformatic Analysis

Cited 2 times

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Genomic DNA was extracted from peripheral blood leukocytes according
to standard procedures, and then exome libraries (Agilent SureSelectXT Reagent
Kit; Agilent Technologies) were sequenced on an Illumina HiSeq 2500 at the
Genomic Technologies Facility in Lausanne, Switzerland. Bioinformatic analyses
were performed as described previously.^{5 (link)} Briefly, raw reads were
mapped to the human reference genome (hg19/GRCh37) using the Novoalign software
(V3.08.00, Novocraft Technologies). Next, Picard (version 2.14.0-SNAPSHOT) was
used to remove duplicate reads and Genome Analysis Toolkit (GATK) (version
3.8)^{6 (link)} was used to perform base quality score
recalibration on both single-nucleotide variants and insertion–deletions. A VCF
file with the variants was generated by HaplotypeCaller. Then, DNA variants were
filtered based on quality, frequency in ExAC, gnomAD,^{7 (link)} 1000
Genomes,^{8 (link)} ESP (NHLBI Exome Variant Server, http://evs.gs.washington.edu/EVS), GME (GME Variome http://igm.ucsd.edu/gme/index.php), and ABraOM,^{9 (link)} and on predicted impact on protein
sequence and messenger RNA (mRNA) splicing. Finally, they were annotated
according to a specific in-house pipeline.^{5 (link)}

Peter V.G., Quinodoz M., Pinto-Basto J., Sousa S.B., Di Gioia S.A., Soares G., Ferraz Leal G., Silva E.D., Pescini Gobert R., Miyake N., Matsumoto N., Engle E.C., Unger S., Shapiro F., Superti-Furga A., Rivolta C, & Campos-Xavier B. (2019). The Liberfarb syndrome, a multisystem disorder affecting eye, ear, bone, and brain development, is caused by a founder pathogenic variant in the PISD gene. Genetics in Medicine, 21(12), 2734-2743.

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Targeted Exome Sequencing Protocol

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Genomic DNA was fragmented to a size range of 150–250 bp using the Covaris S220 instrument. The fragmented DNA was end repaired and adenylated. The SureSelect Adaptor was then ligated followed by a pre-amplification. After library preparation, target regions were captured by hybridization of biotinylated baits (library probes) using Agilent SureSelect XT Human All Exon V6 (Agilent, Santa Clara, CA, USA). Captured target sequences were then isolated using streptavidin-coated magnetic beads. Subsequently, the appropriate 8-bp single index tags were added during sequencing library amplification. All steps were done using Agilent SureSelect XT Reagent Kit (Agilent, Santa Clara, CA, USA). Final quality control was done using Qubit 3 fluorometer and Agilent Bioanalyzer 2100 (Agilent, Santa Clara, CA, USA). The libraries were sequenced in paired-end mode (2 × 50 nt) on a NovaSeq6000 S2 Flowcell (Illumina, San Diego, CA, USA) resulting in ~200 million distinct sequencing reads per library.

Costa B., Fletcher M.N., Boskovic P., Ivanova E.L., Eisemann T., Lohr S., Bunse L., Löwer M., Burchard S., Korshunov A., Coltella N., Cusimano M., Naldini L., Liu H.K., Platten M., Radlwimmer B., Angel P, & Peterziel H. (2021). A Set of Cell Lines Derived from a Genetic Murine Glioblastoma Model Recapitulates Molecular and Morphological Characteristics of Human Tumors. Cancers, 13(2), 230.

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Exome Sequencing of Peripheral Blood Samples

Cited 2 times

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Genomic DNA was extracted from peripheral blood leukocytes according to standard procedures, and then exome libraries (Agilent SureSelectXT Reagent Kit; Agilent Technologies) were sequenced on an Illumina HiSeq 2500 at the Genomic Technologies Facility in Lausanne, Switzerland. Bioinformatic analyses were performed as described before.^{5 (link)} Briefly, raw reads were mapped to the human reference genome (hg19/GRCh37) using the Novoalign software (V3.08.00, Novocraft Technologies). Next, Picard (version 2.14.0-SNAPSHOT) was used to remove duplicate reads and GATK was used (version 3.8)^{6 (link)} to perform ‘base quality score recalibration’ on both single nucleotide variants and insertion-deletions. A VCF file with the variants was generated by HaplotypeCaller. Then, DNA variants were filtered based on quality, frequency in ExAC, gnomAD,^{7 (link)} 1000 Genomes,^{8 (link)} ESP (NHLBI Exome Variant Server, http://evs.gs.washington.edu/EVS), GME (GME Variome http://igm.ucsd.edu/gme/index.php) and ABraOM,^{9 (link)} and on predicted impact on protein sequence and mRNA splicing. Finally, they were annotated according to a specific in-house pipeline.^{5 (link)}

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Whole Exome Sequencing Library Preparation

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Two hundred ng of genomic DNA were used for library preparation, using the Agilent SureSelectXT reagent kit (Catalog number G9642B, Agilent Technologies, Inc.)and the All Exon v5 probeset (5190-8863, Agilent Technologies, Inc.). Following hybridization, the libraries were purified according to the manufacturer's recommendations and amplified by polymerase chain reaction (12 cycles). DNA integrity was verified using TapeStation (SCREENTAPE D1000 tapestation 5067-5582, reagents D1000 tapestation 5067-5583). Concentrations were measured using Qubit^® dsDNA BR Assay Q32853. Loading concentrations were 22 nM for fragmentation and 6 pM for NextSeq injection. Normalized libraries were pooled, and DNA was sequenced on an Illumina NextSeq500 device using 2x111-bp paired-end reads and multiplexed. Names, catalog numbers and suppliers of the Illumina sequencing kit were following: NextSeq 500 High Output Kit FC404-2004/2140817 and NextSeq 500 Mid Output Kit FC404-2003/2140816. More than 90% of the target sequence was covered with a read depth of at least 10X for somatic DNA.

D'Agay M.D., Galland L., Tharin Z., Truntzer C, & Ghiringhelli F. (2021). Utility of exome sequencing in routine care for metastatic colorectal cancer. Molecular and Clinical Oncology, 15(5), 229.

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Whole Exome Sequencing of Primary and Metastatic Tumors

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PIM1-CBRLuc primary MFP tumors and lung metastases were analyzed by WES performed by the Sequencing and Microarray Facility at MD Anderson Cancer Center. Libraries were prepared from 200 ng of Biorupter ultrasonicator (Diagenode)-sheared gDNA using the Agilent SureSelectXT Reagent Kit (Agilent Technologies). Libraries were prepared for capture with ten cycles of PCR amplification, then assessed for size distribution on a Fragment Analyzer using the High Sensitivity NGS Fragment Analysis Kit (Advanced Analyticals) and quantified using the Qubit dsDNA HS Assay Kit (ThermoFisher). Exon target capture was performed using the Agilent SureSelectXT Human All Exon V4 kit. Following capture, index tags were added to the exon enriched libraries using six cycles of PCR. The indexed libraries were then assessed for size distribution using the Agilent TapeStation and quantified using the Qubit dsDNA HS Assay Kit. Libraries were sequenced either one library per lane (5 samples) or equal molar concentrations of 3 libraries were pooled and sequenced in two lanes of the HiSeq4000 sequencer, using the 75nt paired end format.

Echeverria G.V., Powell E., Seth S., Ge Z., Carugo A., Bristow C., Peoples M., Robinson F., Qiu H., Shao J., Jeter-Jones S.L., Zhang X., Ramamoorthy V., Cai S., Wu W., Draetta G., Moulder S.L., Symmans W.F., Chang J.T., Heffernan T.P, & Piwnica-Worms H. (2018). High-resolution clonal mapping of multi-organ metastasis in triple negative breast cancer. Nature Communications, 9, 5079.

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Agilent SureSelectXT Exome Sequencing

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Two hundred ng of genomic DNA were used for library preparation, using the Agilent SureSelectXT reagent kit (Catalog number G9642B, Agilent Technologies, Inc., Santa Clara, CA, USA) and the All Exon v5 probeset (5190–8863, Agilent Technologies, Inc.). Following hybridization, the libraries were purified according to the manufacturer’s recommendations and amplified by polymerase chain reaction (12 cycles). DNA integrity was verified using TapeStation (SCREENTAPE D1000 tapestation 5067–5582, reagents D1000 tapestation 5067–5583). Concentrations were measured using Qubit^® dsDNA BR Assay Q32853 (Thermo Fisher Scientific, Waltham, MA, USA). Loading concentrations were 22 nM for fragmentation and 6 pM for NextSeq.

Lecuelle J., Boidot R., Mananet H., Derangère V., Albuisson J., Goussot V., Arnould L., Tharin Z., Ray Coquard I., Ghiringhelli F., Truntzer C, & Fumet J.D. (2021). TCR Clonality and Genomic Instability Signatures as Prognostic Biomarkers in High Grade Serous Ovarian Cancer. Cancers, 13(17), 4394.

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TP53 Mutation Detection by Targeted Sequencing

Cited 1 time

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The targeted massive parallel sequencing of the tumour DNA generated data on the TP53 mutation status. The fragmentation of 1,000 ng dsDNA was achieved using the Covaris® M220 Focused-ultrasonicator™ (Covaris, Woburn, MA, USA). The library preparation was performed using the Agilent SureSelectXT reagent kit (Agilent Technologies, Santa Clara, CA, USA), and the individual samples were run on a MiSeq instrument (Illumina, San Diego, CA, USA). This design included +/−10 nucleotides at exon-intron borders, to cover potential splice site mutations. The TP53 data was extracted from a sequencing effort applying baits targeting 360 genes as previously described in detail^{37 (link)}. Preliminary mutation calling was performed using the MiSeq Reporter (MSR) software, and the raw mutation calling output was revised by the application of post-processing filters. All of the suspected TP53 mutations were validated by the manual inspection of the sequencing reads using the Integrative Genomics Viewer^{38 (link)}.

Bischof K., Knappskog S., Hjelle S.M., Stefansson I., Woie K., Salvesen H.B., Gjertsen B.T, & Bjorge L. (2019). Influence of p53 Isoform Expression on Survival in High-Grade Serous Ovarian Cancers. Scientific Reports, 9, 5244.

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Whole Exome Sequencing Protocol

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Two hundred ng of genomic DNA were used for library preparation, using the Agilent SureSelectXT reagent kit (Agilent Technologies, Santa Clara, CA, USA). The totality of the enriched library was used in the hybridization and captured with the SureSelect All Exon v5 or v6 (Agilent Technologies) baits. Following hybridization, the captured libraries were purified according to the manufacturer’s recommendations and amplified by polymerase chain reaction (12 cycles). Normalized libraries were pooled, and DNA was sequenced on an Illumina NextSeq500 device using 2 ×111-bp paired-end reads and multiplexed. More than 90% of the target sequence was covered with a read depth of at least 10X for somatic DNA (Supplementary Table 3).

Galland L., Ballot E., Mananet H., Boidot R., Lecuelle J., Albuisson J., Arnould L., Desmoulins I., Mayeur D., Kaderbhai C., Ilie S., Hennequin A., Bergeron A., Derangère V., Ghiringhelli F., Truntzer C, & Ladoire S. (2022). Efficacy of platinum-based chemotherapy in metastatic breast cancer and HRD biomarkers: utility of exome sequencing. NPJ Breast Cancer, 8, 28.

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Exome Sequencing of Tumor and Germline DNA

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Two hundred ng of genomic DNA were used for library preparation, using the Agilent SureSelectXT reagent kit (Agilent Technologies, Santa Clara, USA). The totality of enriched library was used in the hybridization and captured with the SureSelect All Exon v5 or v6 (Agilent Technologies) baits. Following hybridization, the captured libraries were purified according to the manufacturer's recommendations and amplified by polymerase chain reaction (12 cycles). Normalized libraries were pooled and DNA was sequenced on an Illumina NextSeq500 device using 2 × 111-bp paired-end reads and multiplexed. Tumor and germline DNA sequencing generated mean target coverages of 78X and 90X respectively, and a mean of more than 90% of the target sequence was covered with a read depth of at least 10X for somatic DNA.

Réda M., Richard C., Bertaut A., Niogret J., Collot T., Fumet J.D., Blanc J., Truntzer C., Desmoulins I., Ladoire S., Hennequin A., Favier L., Bengrine L., Vincent J., Hervieu A., Dusserre J.F., Lepage C., Foucher P., Borg C., Albuisson J., Arnould L., Nambot S., Faivre L., Boidot R, & Ghiringhelli F. (2020). Implementation and use of whole exome sequencing for metastatic solid cancer. EBioMedicine, 51, 102624.

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Exome Sequencing Library Preparation

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Two hundred ng of genomic DNA was used for library preparation, using the Agilent SureSelectXT reagent kit (Agilent Technologies, Santa Clara, CA, USA). The totality of the enriched library was used in the hybridization and captured with the SureSelect All Exon v5 or v6 (Agilent Technologies) baits. Following hybridization, the captured libraries were purified according to the manufacturer’s recommendations and amplified by polymerase chain reaction (12 cycles). Normalized libraries were pooled, and DNA was sequenced on an Illumina NextSeq500 device using 2 × 111 bp paired-end reads and multiplexed. Tumor and germline DNA sequencing generated mean target coverages of 78× and 90×, respectively, and a mean of more than 90% of the target sequence was covered with a read depth of at least 10× for somatic DNA.

Dalens L., Lecuelle J., Favier L., Fraisse C., Lagrange A., Kaderbhai C., Boidot R., Chevrier S., Mananet H., Derangère V., Truntzer C, & Ghiringhelli F. (2023). Exome-Based Genomic Markers Could Improve Prediction of Checkpoint Inhibitor Efficacy Independently of Tumor Type. International Journal of Molecular Sciences, 24(8), 7592.

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