A total of 504 cancer-related genes were presented in Supplementary Table 2 , and targeted for capture and deep sequencing. Using the eArray system (Agilent Technologies, Santa Clara, CA), the capture was designed to include all of protein coding sequences (CDSs) and most of the untranslated regions of these genes. In accordance with the manufacturer's protocol, genomic DNA was fragmented by the NEBNext dsDNA Fragmentase (New England Biolabs, Ipswich, MA), and daptor-ligated library was constructed using a Agilent SureSelect library kit (Agilent Technologies, Santa Clara, CA). Targeted sequence enrichment was performed using the Agilent SureSelect Target Enrichment Kit (Agilent Technologies, Santa Clara, CA) according to the manufacturer's instructions. The enriched samples were sequenced via 2 × 100 paried-end sequencing using a Hiseq 2000 Sequencing system (Illumina, San Diego, CA). Illumina Sequencing Control v2.8, Illumina Off-Line Basecaller v1.8 and Illumina Consensus Assessment of Sequence and Variation v1.8 software (Illumina, San Diego, CA) were used to produce 100 bp sequence reads.
Earray system
Manufactured by Agilent Technologies
Sourced in United States
The EArray system is a versatile and powerful platform for DNA microarray analysis. It offers a comprehensive solution for DNA microarray experimentation, including array design, sample preparation, hybridization, and data analysis. The EArray system provides researchers with the tools to conduct high-throughput genomic studies, enabling them to explore gene expression, genotyping, and DNA methylation patterns.
Automatically generated - may contain errors
Lab products found in correlation
Off line basecaller v1, by Illumina (2 mentions)
Nebnext dsdna fragmentase, by New England Biolabs (2 mentions)
Agilent sureselect target enrichment kit, by Agilent Technologies (2 mentions)
Hiseq 2000 sequencing system, by Illumina (2 mentions)
Sequencing control v2, by Illumina (2 mentions)
Consensus assessment of sequence and variation v1, by Illumina (2 mentions)
Sureprint g3 array, by Agilent Technologies (1 mentions)
Dneasy blood tissue kit, by Qiagen (1 mentions)
Paired end genomic dna kits, by Illumina (1 mentions)
Hiseq platform, by Illumina (1 mentions)
S1 ultrasonicator, by Covaris (1 mentions)
Feature extraction software, by Agilent Technologies (1 mentions)
Low input quick amp wt labeling kit, by Agilent Technologies (1 mentions)
9 protocols using earray system
1
Cancer Gene Sequencing Protocol
2
Microarray Analysis of Differential Gene Expression
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Custom-designed 8-by-60,000 SurePrint G3 arrays from Agilent Technologies were used for microarray analysis. The arrays were designed to cover at least two independent probes per gene as well as the intergenic regions of the UTI89 genome in four replicates and 1,319 control probes for detection of hybridization efficiency as well as eukaryotic control probes, using the eArray system (Agilent Technologies). Hybridization was carried out using a gene expression hybridization kit and wash buffers (Agilent Technologies) as per the manufacturer’s recommendations and scanned using an Agilent scanner. Feature-extracted files were analyzed using DNAStar software. Probe signal intensities were subjected to background correction, quantile normalization, and median polish. To identify differentially expressed transcripts, the log2 fold change was calculated (cutoff, ≥2 log2 fold change), and filtering was based on Student’s t test (significance cutoff, P < 0.05). The statistically significant, differentially expressed transcripts were further examined based on gene ontology information obtained from EcoCyc and literature searches.
3
Pathogen-Specific Microarray for P. salmonis
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In order to analyze P. salmonis gene expression, we designed a pathogen-specific oligo microarray using the eArray system (Agilent Technologies). P. salmonis LF-89 gene targets were derived from the genome assembly and annotation was reported in Pulgar et al. (2015b (link)). Each array contained 15,208 60-mer oligonucleotides (5310 different probes) representing 2850 P. salmonis gene targets with at least two duplicate probes per gene. The array also contained positive and negative controls, previously selected by Agilent for use in commercial microarrays. EST sequences of Salmo salar were used as queries in the design step to reduce the likelihood of designing probes that would cross-hybridize with host sequences. Selected probes were synthesized in situ on a glass slide in an 8 × 15 K format using Agilent SurePrint technology. Specificity and accuracy of the microarray for bacterial transcripts were assessed by performing hybridizations with gDNA from SHK-1 and from P. salmonis (data not shown). gDNA was isolated using the DNeasy Blood & Tissue kit (Qiagen) from T-25 flasks of confluent SHK-1 cells or 1 ml of exponential phase P. salmonis culture, according to the manufacturer's instructions.
4
High-Resolution aCGH for Neuropathy Gene CNV
5
Targeted Sequencing of Neurological Disorder Genes
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A total of 355 genes were targeted for capture and deep sequencing. They include candidate genes associated with DD/ID, microdeletion/microduplication syndromes, congenital anomalies, and autism spectrum disorders (Table S1 ). Using the eArray system (Agilent Technologies Inc. USA), the capture was designed to include exons with at least 30 bp of the flanking intronic sequence. In addition, 5 kb of the flanking sequence in the 5′UTR and 3′UTR regions were added to the targeted regions. A total of 4150 regions from the 355 genes were targeted for a final capture size of 4.79 Mb.
Targeted sequence enrichment was performed using the Agilent SureSelect Target Enrichment Kit (Agilent Technologies Inc. USA). Genomic DNA was sheared using a Covaris S1 Ultrasonicator (Covaris, MA). Adaptor-ligated libraries were constructed using Paired-End Genomic DNA kits (Illumina, CA). The multiplexed samples were sequenced on the Illumina Hiseq platform using 76-bp paired-end reads.
Targeted sequence enrichment was performed using the Agilent SureSelect Target Enrichment Kit (Agilent Technologies Inc. USA). Genomic DNA was sheared using a Covaris S1 Ultrasonicator (Covaris, MA). Adaptor-ligated libraries were constructed using Paired-End Genomic DNA kits (Illumina, CA). The multiplexed samples were sequenced on the Illumina Hiseq platform using 76-bp paired-end reads.
6
Comprehensive Cancer Gene Panel Sequencing
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A total of 504 cancer-related genes were targeted for capture and deep sequencing. Using the eArray system (Agilent, CA), the capture was designed to include all of protein coding sequences and most of the untranslated regions of these genes. In accordance with the manufacturer’s protocol, genomic DNA was fragmented by the NEBNext dsDNA Fragmentase (New England Biolabs, MA) and adaptor-ligated library was constructed using an Agilent SureSelect library kit (Agilent, CA). Targeted sequence enrichment was performed using the Agilent SureSelect Target Enrichment Kit (Agilent, CA) according to the manufacturer’s instructions. The enriched samples were sequenced via 2 × 100 paired-end sequencing using a HiSeq2000 Sequencing System (Illumina, CA). Illumina Sequencing Control v2.8, Illumina Off-Line Basecaller v1.8, and Illumina Consensus Assessment of Sequence and Variation v1.8 software (Illumina, CA) were used to produce 100 bp sequence reads.
7
Differentially Expressed Genes Analysis
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A custom oligonucleotide spotted microarray (ID: 074275) based on the genome sequence of LcS was designed, produced, and validated by using the eArray system (Agilent Technologies). Cyanine-3-labeled cRNA was obtained from 100 ng of extracted total RNA by using a Low Input Quick Amp WT Labeling Kit (Agilent Technologies). Hybridization was performed on custom microarrays at 65°C for 17 h by using the prepared cRNA. The slides were then washed, and images of the microarrays were acquired by using an Agilent G4900DA SureScan Scanner (Agilent Technologies). The signal data for each spot were subsequently quantified by using Feature Extraction software (Agilent Technologies). The limma package (v.3.36.5) (51 (link)) of the statistical analysis software R (v4.2.1) was used for background correction, quantile normalization of the probe signal data, and identification of differentially expressed genes. The NCBI’s Conserved Domain Search was utilized for the COG classification of genes. Enrichment analysis of COGs was performed by using the clusterProfiler package (v4.4.4) (52 (link)).
8
Custom Gene Expression Microarray for Locust Transcriptomics
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A 2 x 105K Custom Gene Expression Microarray slide (Agilent) was spotted with probes that represented 12107 unigenes from LocustDB (Ma et al., 2006) (link), all L. migratoria ESTs from the CNS deep-sequencing project (Zhang et al., 2012) (link) that did not appear to be represented in LocustDB, and all S. gregaria EST sequences (Badisco et al., 2011) (link) that had no ortholog in the L. migratoria (blast-N hits producing E-value < 1e-10 were considered as orthologs). For each of the resulting 48802 unique sequences, two different probes were spotted in addition to standard Agilent control features.
Probes were designed by means of the eArray system (Agilent). The best probe methodology was used selecting two probes per target, with a probe length of 60 nucleotides in sense orientation. The remainder of the array was supplemented with random doubles, which acted as technical replicates in the analyses. The data and platform information have been deposited in NCBI's Gene Expression Omnibus (Edgar et al., 2002) (link) and are accessible through GEO Series accession number GSE83967 (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE83967).
Probes were designed by means of the eArray system (Agilent). The best probe methodology was used selecting two probes per target, with a probe length of 60 nucleotides in sense orientation. The remainder of the array was supplemented with random doubles, which acted as technical replicates in the analyses. The data and platform information have been deposited in NCBI's Gene Expression Omnibus (Edgar et al., 2002) (link) and are accessible through GEO Series accession number GSE83967 (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE83967).
9
Custom aCGH for Genome-Wide CNV Detection
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A custom aCGH was developed to detect CNV on the human genome. First, genomic sequences (hg19) of the whole Y chromosome, AR gene on chromosome X, CFTR gene on chromosome 7, PLOG gene on chromosome 15, INSL3 gene on chromosome 19, RXFP1 gene on chromosome 4 and mitochondrion were downloaded from NCBI (http:// www.ncbi.nlm.nih.gov/). Then, 60-mer probes with matched melting temperature were selected to remove those probes with secondary genomic alignments on the genome. After the most stringent filter-ing, 56,205 probes were generated. Specifically, there were 50,615 probes on chromosome Y, 1694 probes covering AR gene on chromosome X, 2124 probes covering CFTR gene on chromosome 7, 253 probes covering POLG gene on chromosome 15, 527 probes covering INSL3 gene on chromosome 19, 1161 probes covering RXFP1 gene on chromosome 4 and 1,331 probes covering mitochondrial DNA. All of the probes with average probe spacing of 500 bp were submitted to Agilent's E-array system (Agilent, Santa Clara, CA, USA) to produce the custom array.
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