Agilent sureselect v 4 kit

Manufactured by Agilent Technologies

Sourced in United States

The Agilent SureSelect v.4 kit is a targeted enrichment solution for next-generation sequencing. It allows for the selective capture and sequencing of specific regions of the genome, enabling in-depth analysis of genomic targets of interest.

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Lab products found in correlation

Truseq library construction, by Illumina (4 mentions) Hiseq 2000 2500, by Illumina (3 mentions) Dna blood mini kit, by Qiagen (2 mentions) Dna ffpe, by Qiagen (2 mentions) Hiseq system, by Illumina (2 mentions) Dneasy blood and tissue mini kit, by Qiagen (1 mentions) Dna kit, by Qiagen (1 mentions) Hiseq 2500, by Illumina (1 mentions) Hiseq 2500 platform, by Illumina (1 mentions)

8 protocols using agilent sureselect v 4 kit

Targeted Exome Sequencing of Tumor and Normal

Cited 2 times

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Samples were prepared, sequenced and analyzed as described earlier (Bertotti et al., 2015 (link)). DNA was extracted from cells and xenograft tissues using the Qiagen DNeasy blood and tissue mini kit (Qiagen, CA). Fragmented genomic DNA from tumor and normal samples was used for Illumina TruSeq library construction (Illumina, San Diego, CA). Exonic regions or targeted regions (targeted genes-Table S3) were captured in solution using the Agilent SureSelect v.4 kit (Agilent, Santa Clara, CA) according to the manufacturer’s instructions as previously described (Bertotti et al., 2015 (link); Sausen et al., 2013 (link)). Paired-end sequencing, resulting in 100 or 150 bases from each end of the DNA fragments for the exome or targeted sequencing libraries, respectively, was performed using Illumina HiSeq 2000/2500 instrumentation (Illumina, San Diego, CA).

Vaz M., Hwang S.Y., Kagiampakis I., Phallen J., Patil A., O’Hagan H.M., Murphy L., Zahnow C.A., Gabrielson E., Velculescu V.E., Easwaran H.P, & Baylin S.B. (2017). Chronic Cigarette Smoke-Induced Epigenomic Changes Precede Sensitization of Bronchial Epithelial Cells to Single Step Transformation by KRAS Mutations. Cancer cell, 32(3), 360-376.e6.

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Comparative Analysis of Exome Sequencing Techniques

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Exome sequencing samples were collected for two current mainstream technologies. We selected 2 × 12 exome libraries captured with the Agilent SureSelect V4 kit (Agilent, Santa Clara, CA) sequenced by the Beijing Genomics Institute (BGI) in Copenhagen on an Illumina HiSeq system (Illumina, San Diego, CA) with 101 bp paired‐end reads. The first set contained samples sequenced to an average coverage of 78x and the second set were different samples sequenced at 160x. An additional 12 libraries were captured with the latest Agilent SureSelect V5 capture kit and sequenced by the Charité university clinic Berlin to an average coverage of 100x on an Illumina HiSeq system using 100 bp paired‐end reads. For NimbleGen, we selected 12 libraries captured by the latest NimbleGen SeqCap V3 and sequenced on an Illumina HiSeq using 101 bp paired‐end reads at 95x average coverage at the Duke Genome Centre (Table 1). DNA from all samples was derived from blood.

Lelieveld S.H., Spielmann M., Mundlos S., Veltman J.A, & Gilissen C. (2015). Comparison of Exome and Genome Sequencing Technologies for the Complete Capture of Protein‐Coding Regions. Human Mutation, 36(8), 815-822.

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Whole Exome Sequencing of Tumor and Normal Samples

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Whole exome sequencing was performed on pre-treatment tumor and matched normal samples. DNA was extracted from patients’ tumors and matched peripheral blood using the QIAGEN DNA kit (QIAGEN, CA). Fragmented genomic DNA from tumor and normal samples was used for Illumina TruSeq library construction (Illumina, San Diego, CA) and exonic regions were captured in solution using the Agilent SureSelect v.4 kit (Agilent, Santa Clara, CA) according to the manufacturers’ instructions as previously described¹⁰ (link)^,⁴⁷ . Paired-end sequencing, resulting in 100 bases from each end of the fragments for the exome libraries was performed using Illumina HiSeq 2000/2500 instrumentation (Illumina, San Diego, CA). The mean depth of total and distinct coverage for the pre-treatment tumors were 206x and 173x (Table S1O).

Anagnostou V., Bruhm D.C., Niknafs N., White J.R., Shao X.M., Sidhom J.W., Stein J., Tsai H.L., Wang H., Belcaid Z., Murray J., Balan A., Ferreira L., Ross-Macdonald P., Wind-Rotolo M., Baras A.S., Taube J., Karchin R., Scharpf R.B., Grasso C., Ribas A., Pardoll D.M., Topalian S.L, & Velculescu V.E. (2020). Integrative Tumor and Immune Cell Multi-omic Analyses Predict Response to Immune Checkpoint Blockade in Melanoma. Cell Reports Medicine, 1(8), 100139.

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Whole Exome Sequencing of Lung Cancer

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Whole exome sequencing was performed on the pre-treatment and post-progression tumor and matched normal samples. Tumor samples underwent pathological review for confirmation of lung cancer diagnosis and assessment of tumor purity. Slides from each FFPE block were macrodissected to remove contaminating normal tissue. Matched normal samples were provided as peripheral blood. DNA was extracted from patients’ tumors and matched peripheral blood using the Qiagen DNA FFPE and Qiagen DNA blood mini kit respectively (Qiagen, CA). Fragmented genomic DNA from tumor and normal samples was used for Illumina TruSeq library construction (Illumina, San Diego, CA) and exonic regions were captured in solution using the Agilent SureSelect v.4 kit (Agilent, Santa Clara, CA) according to the manufacturers’ instructions as previously described (16 (link), 23 (link), 35 (link)). Paired-end sequencing, resulting in 100 bases from each end of the fragments for the exome libraries was performed using Illumina HiSeq 2000/2500 instrumentation (Illumina, San Diego, CA). The mean depth of coverage for the pre-treatment and resistant tumors was 214x and 217x respectively, allowing us to identify sequence alterations and copy number changes in >20,000 genes (Supplementary Table S2).

Anagnostou V., Smith K.N., Forde P.M., Niknafs N., Bhattacharya R., White J., Zhang T., Adleff V., Phallen J., Wali N., Hruban C., Guthrie V.B., Rodgers K., Naidoo J., Kang H., Sharfman W., Georgiades C., Verde F., Illei P., Li Q.K., Gabrielson E., Brock M.V., Zahnow C.A., Baylin S.B., Scharpf R., Brahmer J.R., Karchin R., Pardoll D.M, & Velculescu V.E. (2016). Evolution of neoantigen landscape during immune checkpoint blockade in non-small cell lung cancer. Cancer discovery, 7(3), 264-276.

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Whole-Exome Sequencing and Variant Filtering

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After obtaining written informed consent, whole‐exome sequencing was done using DNA isolated from blood, as described previously (Lelieveld et al., 2016). Briefly, exome capture was done using the Agilent SureSelect v4 Kit (Agilent, Santa Clara, CA). Exome libraries were sequenced on an Illumina HiSeq instrument (Illumina, San Diego, CA) with 101‐bp paired‐end reads at a median coverage of 75× and with >95% of exons having coverage >30×. Sequence reads were aligned to the hg19 reference genome using BWA version 0.5.9‐r16. Variants were subsequently called by the GATK unified genotyper, version 3.2‐2 and annotated using a custom diagnostic annotation pipeline. Variants were filtered for having less than 1% frequency in dbSNP, having <1% frequency in our in‐house database and having <1% frequency in the “Exome Aggregation Consortium” (ExAC) database (http://www.exac.broadinstitute.org).

Kerkhofs C., Stevens S.J., Faust S.N., Rae W., Williams A.P., Wurm P., Østern R., Fockens P., Würfel C., Laass M., Kokke F., Stegmann A.P, & Brunner H.G. (2019). Mutations in RPSA and NKX2‐3 link development of the spleen and intestinal vasculature. Human Mutation, 41(1), 196-202.

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Trio-Based Whole Exome Sequencing

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For whole exome sequencing (WES), a parent–offspring trio approach was used as described by us previously [3 (link), 4 (link)]. Exomes were sequenced using DNA isolated from blood according to standard procedures. Exome capture was done using the Agilent SureSelect v4 kit (Agilent, Santa Clara, CA, USA). Exome libraries were sequenced on an Illumina HiSeq instrument (Illumina, San Diego, CA, USA) with 101 bp paired-end reads at a median coverage of 75x. Sequence reads were aligned to the hg19 reference genome using BWA version 0.5.9-r16. Variants were subsequently called by the GATK unified genotyper, version 3.2-2 and annotated using a custom diagnostic annotation pipeline. De novo variants in index patients were called as described by de Ligt et al. [4 (link)]. Standard Sanger sequencing of patient and parental DNA was used for validation de novo variants identified in WES data.

Stevens S.J., van Essen A.J., van Ravenswaaij C.M., Elias A.F., Haven J.A., Lelieveld S.H., Pfundt R., Nillesen W.M., Yntema H.G., van Roozendaal K., Stegmann A.P., Gilissen C, & Brunner H.G. (2016). Truncating de novo mutations in the Krüppel-type zinc-finger gene ZNF148 in patients with corpus callosum defects, developmental delay, short stature, and dysmorphisms. Genome Medicine, 8, 131.

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Whole-Exome Sequencing of Lung Cancer

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Whole-exome sequencing was performed on preimmunotherapy tumor and matched normal samples, with the exception of three cases for which tumor from the time of resistance to therapy was analyzed (Supplementary Table 1). Tumor formalin-fixed paraffin-embedded (FFPE) blocks were retrieved and underwent pathological review for confirmation of lung cancer diagnosis and assessment of tumor cellularity. Slides from each FFPE block were macrodissected to remove contaminating normal tissue. Matched normal samples were provided as peripheral blood. DNA was extracted from patients’ tumors and matched peripheral blood using the Qiagen DNA FFPE and Qiagen DNA blood mini kit, respectively (Qiagen). Fragmented genomic DNA from tumor and normal samples used for Illumina TruSeq library construction (Illumina) and exonic regions were captured in solution using the Agilent SureSelect v.4 kit (Agilent) as previously described¹³. Paired-end sequencing, resulting in 100 bases from each end of the fragments for the exome libraries was performed using Illumina HiSeq 2500 instrumentation (Illumina). The mean depth of total and distinct coverage for the pretreatment tumors were ×231 and ×144, respectively (Supplementary Tables 2, 3 and 6).

Anagnostou V., Niknafs N., Marrone K., Bruhm D.C., White J.R., Naidoo J., Hummelink K., Monkhorst K., Lalezari F., Lanis M., Rosner S., Reuss J.E., Smith K.N., Adleff V., Rodgers K., Belcaid Z., Rhymee L., Levy B., Feliciano J., Hann C.L., Ettinger D.S., Georgiades C., Verde F., Illei P., Li Q.K., Baras A.S., Gabrielson E., Brock M.V., Karchin R., Pardoll D.M., Baylin S.B., Brahmer J.R., Scharpf R.B., Forde P.M, & Velculescu V.E. (2020). Multimodal genomic features predict outcome of immune checkpoint blockade in non-small-cell lung cancer. Nature cancer, 1(1), 99-111.

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Whole Exome Sequencing of HSCR

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In a trio design, patients with HSCR and their non-affected parents were analyzed by WES. The Agilent SureSelect V4 kit (Agilent Technologies, Santa Clara, USA) was used to prepare the sequencing library. Sequencing was performed on the Illumina HiSeq 2500 platform (Illumina, San Diego, USA) as described in the Supplementary (S1 Text). Filtered variants for both patients (patient I: 11 genes, patient II: 14 genes) are shown in S1 Table and S2 Table.

Mederer T., Schmitteckert S., Volz J., Martínez C., Röth R., Thumberger T., Eckstein V., Scheuerer J., Thöni C., Lasitschka F., Carstensen L., Günther P., Holland-Cunz S., Hofstra R., Brosens E., Rosenfeld J.A., Schaaf C.P., Schriemer D., Ceccherini I., Rusmini M., Tilghman J., Luzón-Toro B., Torroglosa A., Borrego S., Sze-man Tang C., Garcia-Barceló M., Tam P., Paramasivam N., Bewerunge-Hudler M., De La Torre C., Gretz N., Rappold G.A., Romero P, & Niesler B. (2020). A complementary study approach unravels novel players in the pathoetiology of Hirschsprung disease. PLoS Genetics, 16(11), e1009106.

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