Hiscansq platform

Manufactured by Illumina

Sourced in United States

The HiScanSQ platform is a DNA sequencing system designed by Illumina. It is capable of generating high-quality sequencing data for a wide range of applications.

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Lab products found in correlation

Nanodrop 2000, by Thermo Fisher Scientific (6 mentions) 2100 bioanalyzer, by Agilent Technologies (4 mentions) Nanodrop 2000 spectrophotometer, by Thermo Fisher Scientific (4 mentions) Casava v1, by Illumina (3 mentions) Truseq rna sample preparation kit v2, by Illumina (3 mentions) Agilent bioanalyzer, by Agilent Technologies (2 mentions) Truseq sbs kit v3, by Illumina (2 mentions) Rneasy plant mini kit, by Qiagen (2 mentions) Truseq rna library prep kit, by Illumina (2 mentions) Dna mini kit, by Qiagen (2 mentions) Picogreen assay, by Thermo Fisher Scientific (2 mentions) Nanodrop 1000 spectrophotometer, by Thermo Fisher Scientific (2 mentions) Trueseq exome enrichment kit, by Illumina (1 mentions) Miseq platform, by Illumina (1 mentions) Nextera rapid capture enrichment protocol, by Illumina (1 mentions) Truseq sbs kits v3, by Illumina (1 mentions) Truseq rna sample preparation kits v2, by Illumina (1 mentions) Scriptseq v2 rna seq library preparation kit, by Illumina (1 mentions) Qubit 2.0 fluorometer, by Thermo Fisher Scientific (1 mentions) Wizard genomic dna purification kit, by Promega (1 mentions)

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HiScanSQ (93 citations)

27 protocols using hiscansq platform

Hybrid Seal Genome Sequencing

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Hybrid DNA was isolated from the pulp of a tooth that had fallen from the specimen in storage using the NucleoMag kit (Macherey-Nagel GmbH & Co., Germany). The same method was used to isolate DNA from Baltic grey seal, Baltic ringed and Saimaa ringed seal muscle tissue samples. All libraries and sequencing were performed at the DNA Sequencing and Genomics Laboratory, Institute of Biotechnology, University of Helsinki, Finland. The hybrid seal genome was sequenced with two separate runs of Illumina HiScanSQ platform and 11 separate runs of Illumina MiSeq platform to approximately 100× raw sequencing coverage (electronic supplementary material, table S1). The base calls in the Illumina HiScanSQ platform were converted into text format (FASTA-format with individual base quality scores) using the CASAVA toolkit (bcl2fastq v. 1.8.3) provided by the manufacturer. The Illumina MiSeq platform employed a primary analysis software MiSeq Reporter post-run that produced the base calls in text format.
For all the Illumina reads, base call accuracy and read length filtering (adapter cut-off) were performed with cutadapt [58 (link)] using minimum accepted base call accuracy of 90% and minimum post-filtering read length of 75 bp. The Weddell seal (Leptonychotes weddellii) draft genome (Broad Institute) was used as a reference genome for the downstream genetic distance analysis.

Savriama Y., Valtonen M., Kammonen J.I., Rastas P., Smolander O.P., Lyyski A., Häkkinen T.J., Corfe I.J., Gerber S., Salazar-Ciudad I., Paulin L., Holm L., Löytynoja A., Auvinen P, & Jernvall J. (2018). Bracketing phenogenotypic limits of mammalian hybridization. Royal Society Open Science, 5(11), 180903.

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Small RNA Sequencing and miRNA Analysis

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Small RNA libraries for micro-RNA sequencing were prepared using the TruSeq Small RNA Library Preparation Kit according to the manufacturer's protocol. The size of the library was checked by using a Fragment Analyzer instrument. Samples were sequenced on an Illumina HiScanSQ platform (Illumina). Sequencing reads were trimmed out of the low-quality bases and adapter clipped with Fastx Toolkit. Filtered sequences were mapped and quantified by using miRDeep2 software^{26 (link)} on the most updated version of the human miRBase database.^{27 (link)} For downstream analysis, miRNAs with RPM <100 in all the samples were filtered out. miRNAs with a fold change of log2(FC) ≤1 and log2(FC) ≥1 were considered as upregulated or downregulated in the different populations, respectively.

Bruno S., Tapparo M., Collino F., Chiabotto G., Deregibus M.C., Soares Lindoso R., Neri F., Kholia S., Giunti S., Wen S., Quesenberry P, & Camussi G. (2017). Renal Regenerative Potential of Different Extracellular Vesicle Populations Derived from Bone Marrow Mesenchymal Stromal Cells. Tissue Engineering. Part A, 23(21-22), 1262-1273.

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Whole Exome Sequencing with Illumina Platform

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The WES experiment was performed on Illumina Hi Scan SQ platform and technology (Illumina, San Diego, CA, USA); the True Seq Exome Enrichment Kit enables the enrichment of the coding portion as well as the adjacent intronic and 5’-3’-untranslated regions.

Cogliati F., Giorgini V., Masciadri M., Bonati M.T., Marchi M., Cracco I., Gentilini D., Peron A., Savini M.N., Spaccini L., Scelsa B., Maitz S., Veneselli E., Prato G., Pintaudi M., Moroni I., Vignoli A., Larizza L, & Russo S. (2019). Pathogenic Variants in STXBP1 and in Genes for GABAa Receptor Subunities Cause Atypical Rett/Rett-like Phenotypes. International Journal of Molecular Sciences, 20(15), 3621.

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Transcriptome Sequencing and PA-Seq Analysis

Cited 2 times

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Transcriptome sequencing was performed on an Illumina HiScanSQ platform at the Genomic Medicine and Bioinformatic Core Facilty of the University of Debrecen. Quality assessment of raw read files was achieved with FastQC v0.10.1. Reads were aligned to the respective host genome (Sus scrofa, assembly: Sscrofa10.2) and subsequently to the PRV genome (KJ717942.1), using Tophat v2.09 [30 (link)]; ambiguous reads were discarded. For PA-Seq, mapping was carried out with Bowtie v2. [31 (link)], followed by peak detection using HOMER in strand-specific mode, with adjustments for the peak qualities of oligo(dT) primed libraries. Peak categories were assigned by using in-house scripts, based on the following criteria: the presence or absence of a PAS in the 50 bp region upstream from the PA site and the presence of at least 2 consecutive adenine mismatches in at least 10 independent reads at the PA site. Annotation and visualization were carried out in the Artemis Genome Browser v15.0.0 [32 (link)] and IGV v2.2 [33 (link)]. GC bias in the alignments was inspected by using the Bioconductor R package. The prediction of canonical and non-canonical PAS was carried out using PolyApred [34 (link)].

Oláh P., Tombácz D., Póka N., Csabai Z., Prazsák I, & Boldogkői Z. (2015). Characterization of pseudorabies virus transcriptome by Illumina sequencing. BMC Microbiology, 15, 130.

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Whole Exome Sequencing Workflow

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WES was performed in the three patients carrying rare NBN variants: 100 ng of genomic DNA extracted from peripheral blood was fragmented according to the Nextera^® Rapid Capture Enrichment protocol (Illumina Inc., San Diego, CA, USA). The fragmentation was verified using the Agilent Bioanalyzer and exomes were captured by hybridization using the coding exome exon kit (Nextera). A total of 500 ng of libraries were sequenced with the Illumina HiScan SQ platform at 100bp paired-ends. After that, reads were aligned with BWA to the reference genome (hg19). Aligned reads were treated for realignment and base quality score recalibration with GATK and for duplicate removal with PicardTools (http://picartools.sourceforge.net). The alignment statistics were collected with SAMtools and GATK. Coverage statistics over the targeted regions were calculated with GATK. Variant calling and filtering by quality was performed by GATK. Variants passing quality filters were annotated separately against NCBI RefGene (http://www.ncbi.nlm.nih.gov) and UCSC KnownGene (http://genome.ucsc.edu).

Zuntini R., Bonora E., Pradella L.M., Amato L.B., Vidone M., De Fanti S., Catucci I., Cortesi L., Medici V., Ferrari S., Gasparre G., Peterlongo P., Sazzini M, & Turchetti D. (2021). Detecting Variants in the NBN Gene While Testing for Hereditary Breast Cancer: What to Do Next?. International Journal of Molecular Sciences, 22(11), 5832.

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Transcriptomic Analysis of Fungal Mycelium

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For RNA-Seq analysis, pooled mycelia from 3 replicated cultures were used to prepare an RNA sample per each strain, growing condition and sampling time. Frozen mycelium (100 mg) was powdered in liquid nitrogen. Total RNA was extracted and purified by using RNeasy Plant Mini Kit (Qiagen, Milan, Italy), following the manufacturer's protocol. RNA quantity and quality were determined with a Nanodrop 2000 spectrophotometer (Thermo Fisher Scientific Inc., Wilmington, Delaware, USA) and a Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, USA). cDNA libraries were prepared from 4 μg total RNA using TruSeq RNA Sample Preparation Kit v2 (Illumina, Inc., San Diego, California, USA) and validated according to Illumina’s low-throughput protocol. After normalization, cDNA libraries were pooled for multiplexing before loading onto a flow cell (8–9 samples per lane). The hybridization and cluster generation were performed on a cBot System using TruSeq SR Cluster Kit v3. Sequencing was performed on an Illumina HiScanSQ platform using TruSeq SBS kit v3 (Illumina, Inc.) to obtain Single Reads, 50 nt in length. Indexed raw sequencing reads from each library were de-multiplexed using the CASAVA v1.8 software (Illumina, Inc.).

Gerin D., De Miccolis Angelini R.M., Pollastro S, & Faretra F. (2016). RNA-Seq Reveals OTA-Related Gene Transcriptional Changes in Aspergillus carbonarius. PLoS ONE, 11(1), e0147089.

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Whole Exome Sequencing in PHTS Patients

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Whole Exome Sequencing (WES) was performed in the patient who presented with multiple melanomas/dysplastic nevus in addition to features suggestive of PHTS and his parents (trio analysis); 100 ng of genomic DNA extracted from peripheral blood were tagmented according to Nextera® Rapid Capture Enrichment protocol. The fragmentation was verified using the Agilent Bioanalyzer and library enrichment was performed by hybridization using the coding exome exon kit (Nextera) and sequenced with Illumina HiScan SQ platform at 100 bp paired-ends. Reads were aligned with BWA to the reference genome (hg19). Aligned reads were treated for realignment and base quality score recalibration with GATK, and for duplicate removal with PicardTools (http://picartools.sourceforge.net). Alignment statistics were collected with SAMtools and GATK. Coverage statistics over the targeted regions were calculated with GATK. Variant calling and filtering by quality was performed by GATK. Variants passing quality filters were annotated separately against NCBI RefGene (http://www.ncbi.nlm.nih.gov) and UCSC KnownGene (http://genome.ucsc.edu).

Innella G., Bonora E., Neri I., Virdi A., Guglielmo A., Pradella L.M., Ceccarelli C., Amato L.B., Lanzoni A., Miccoli S., Gasparre G., Zuntini R, & Turchetti D. (2021). PTEN Hamartoma Tumor Syndrome: Skin Manifestations and Insights Into Their Molecular Pathogenesis. Frontiers in Medicine, 8, 688105.

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Small RNA Sequencing Library Preparation

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Small-RNA libraries were prepared using the NEBNext Multiplex Small RNA Library Prep Set for Illumina (Set 1&2) (New England Biolabs, MA, USA), following the manufacturer’s protocol. Briefly, 5′ and 3′ adapters were ligated with small RNA samples, followed by a cDNA library construction and incorporation of index tags by reverse transcription-PCR (RT-PCR). The products of this RT-PCR were purified using 6% non-denaturing polyacrylamide gel electrophoresis, and a 145-160 bp size fraction was isolated. The cDNA library samples were used for cluster generation and Illumina sequencing on the HiScanSQ platform (50 bp single read).
The first step was to assess the quality of the Illumina raw sequences with the FastQC software. Based on the results obtained, the sequence reads were trimmed to remove sequencing adapters and low quality bases. Once the data were deemed of sufficient quality, they were mapped against the human Hg38 build reference sequence, taken from the UCSC Genome Browser. After that, the intersection between the aligned position of reads and the miRNA coordinates taken from miRBase v21 was performed. The alignment and quantification steps were performed using the Subread^{55 (link)} and RSubread^{56 (link)} packages, respectively.

Seco-Cervera M., González-Rodríguez D., Ibáñez-Cabellos J.S., Peiró-Chova L., González-Cabo P., García-López E., Vílchez J.J., Sanz-Gallego I., Pallardó F.V, & García-Giménez J.L. (2017). Circulating miR-323-3p is a biomarker for cardiomyopathy and an indicator of phenotypic variability in Friedreich’s ataxia patients. Scientific Reports, 7, 5237.

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RNA-Seq Analysis of Fruit Samples

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For the RNA-Seq analysis, the fruit were ground using an homogeniser (Ultra-Turrax T25; Janke and Kunkel IKA-Labortechnik, Staufen, Germany), and the total RNA was extracted from 1 g of the frozen-powder homogenate, according to Landi et al. (2014) (link). The RNA quantity and quality were determined using a Nanodrop 2000 (Thermo Fisher Scientific Inc., Wilmington, DE, USA) and a bioanalyzer (model 2100; Agilent Technologies, Santa Clara, CA, USA). cDNA libraries were prepared from 4 μg total RNA using TruSeq RNA Sample Preparation kits v2 (Illumina, Inc., San Diego, CA, USA), and validated according to the Illumina low-throughput protocol. After normalization, the cDNA libraries were pooled for multiplexing, before loading onto a flow cell (five samples per lane). The hybridization and cluster generation were performed on a cBot System using TruSeq SR Cluster kits v3 (Illumina). The sequencing was performed with an Illumina HiScanSQ platform, using TruSeq SBS kits v3 (Illumina) to obtain single reads 50 nt in length. The indexed raw sequencing reads from each library were de-multiplexed using the CASAVA v1.8 software (Illumina).

Landi L., De Miccolis Angelini R.M., Pollastro S., Feliziani E., Faretra F, & Romanazzi G. (2017). Global Transcriptome Analysis and Identification of Differentially Expressed Genes in Strawberry after Preharvest Application of Benzothiadiazole and Chitosan. Frontiers in Plant Science, 8, 235.

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RNA-seq for Mouse Macrophages

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Libraries for sequencing were prepared using the TruSeq RNA Sample Preparation Kit v2 (Illumina Inc., San Diego, CA) following the manufacturer’s instructions. Single-read 50-nt sequencing of pooled libraries was carried out in a HiScanSQ platform (Illumina Inc.). The data were generated from macrophages differentiated from three independent mice.

Carreras-González A., Navasa N., Martín-Ruiz I., Lavín J.L., Azkargorta M., Atondo E., Barriales D., Macías-Cámara N., Pascual-Itoiz M.A., Sampedro L., Tomás-Cortázar J., Peña-Cearra A., Pellón A., Prados-Rosales R., Abecia L., Elortza F., Aransay A.M., Rodríguez H, & Anguita J. (2018). A multi-omic analysis reveals the regulatory role of CD180 during the response of macrophages to Borrelia burgdorferi. Emerging Microbes & Infections, 7, 19.

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