Nextgene software

Manufactured by SoftGenetics

Sourced in United States

NextGENe software is a bioinformatics tool designed for the analysis of next-generation sequencing (NGS) data. The software provides a comprehensive suite of functionalities for tasks such as sequence alignment, variant calling, and data visualization. It is capable of processing a wide range of NGS data formats and supports multiple genome assemblies.

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Lab products found in correlation

Hiseq 2500, by Illumina (6 mentions) Hiseq 2000 sequencer, by Illumina (6 mentions) Nimblegen seqcap probes, by Roche (5 mentions) Ingenuity variant analysis, by Qiagen (4 mentions) Novaseq 6000 sequencer, by Illumina (4 mentions) Casava version 2, by Illumina (4 mentions) Relaxgene blood dna system, by Tiangen Biotech (4 mentions) Ion torrent personal genome machine, by Thermo Fisher Scientific (3 mentions) Sureselectxt human all exon v5 utrs, by Agilent Technologies (3 mentions) Ion ampliseq library kit2 384, by Thermo Fisher Scientific (2 mentions) Ion pgm, by Thermo Fisher Scientific (2 mentions) Dna rna kit, by Qiagen (2 mentions) V2 kits, by Illumina (2 mentions) Kapa library preparation hyper plus kit, by Roche (2 mentions) Dna sample preparation kit, by Roche (2 mentions) Seqcap ez choice library, by Roche (2 mentions) Kapa hyperchoice max library, by Roche (2 mentions) Sureselect target enrichment system, by Agilent Technologies (2 mentions) Qiaamp dna micro kit, by Qiagen (2 mentions) Ion torrent pgm, by Thermo Fisher Scientific (2 mentions)

Spelling variants of this product

NextGENe (50 citations) NextGENe software package (3 citations) NextGENe Software Suite v3.4.2 (2 citations) NextGene software version 2.4.1 (2 citations)

77 protocols using nextgene software

Analytical Performance Evaluation of Genetic Variants

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Evaluation of analytic sensitivity, analytic specificity, and accuracy was performed with the candidate variants in the ROI which spanned all protein-coding regions and intron-exon boundaries (±20 bp). In the analytical performance analysis, “positive” indicates the case where the variants were detected in both a buffy and tumor samples (FFPE and/or FF) or confirmed by Sanger sequencing. “Negative” means that the variants were not detected in buffy or not confirmed by Sanger sequencing.
In a previous study, we deduced parameters corresponding to CNV analysis using NextGENe software (SoftGenetics, LLC). Similarly, CNV analysis was performed using NextGENe software (SoftGenetics, LLC).
All statistical analyses were performed using MedCalc Software (https://www.medcalc.org/), and p values less than 0.05 were regarded as significant. Mann-Whitney U was used to compare differences between two independent groups that were not normally distributed.

Kim Y., Cho C.H., Ha J.S., Kim D.H., Kwon S.Y., Oh S.C, & Lee K.A. (2019). An optimized BRCA1/2 next-generation sequencing for different clinical sample types. Journal of Gynecologic Oncology, 31(1), e9.

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Targeted Sequencing of CPA1 and CPB1

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DNA was extracted as previously described (10 (link), 11 (link)). DNA was sequenced using an AmpliSeq Custom Panel designed to amplify the coding regions of CPA1 and CPB1. NGS was performed with 540 chips (Ion S5 system, single-read, 200 bp read-length) and reads from the Ion Torrent Server were mapped (HG19) and variants called using NextGENe Software (Softgenetics, LLC, State College, PA) according to manufacturer’s protocols as previously described (38 (link)). Median depth of coverage for CPA1 and CPB1 reads was 262 reads per amplicon. A minimum percentage coverage of reads of ≥10 was required for at least 85% of amplicons. Samples below this coverage were re-sequenced to obtain sufficient coverage. An amplicon depth of coverage of 15 reads was required to call a variant, and candidate variant reads of unknown significance were visualized with Integrated Genomics Viewer (IGV) software. Candidate variants of unknown significance were confirmed by Sanger sequencing, performed at the Johns Hopkins DNA sequencing core.

Kawamoto M., Kohi S., Abe T., Dbouk M., Macgregor-Das A., Koi C., Song K.B., Borges M., Sugimine R., Laheru D., Hruban R.H., Roberts N., Klein A.P, & Goggins M. (2021). Endoplasmic stress-inducing variants in CPB1 and CPA1 and risk of pancreatic cancer; a case-control study and meta-analysis. International journal of cancer, 150(7), 1123-1133.

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Targeted Sequencing of Myeloid Malignancies

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Bone marrow mononuclear cells were purified on Ficoll gradients, and pellets were processed for nucleic acid extraction using a DNA/RNA Kit (Qiagen). Genomic DNA was studied by high-throughput sequencing of 45 genes recurrently mutated in myeloid malignancies using a panel designed on Human genome hg19, and sequencing was performed on Ion PGM™ (Life Technologies) on a dedicated 318 V2 chip [60 ]. Libraries were prepared using Ion AmpliSeq library kit2 384 (Life Technologies) according to the manufacturer’s instructions. The average coverage per gene was ≥ 500×. Reads were aligned against human genome build 19 (hg19) and analyzed for single nucleotide variant (SNV) calling with the NextGENe software (SoftGenetics, Chicago, IL) and with an in-house pipeline (Polydiag, Institut Imagine, Université de Paris). We reported all clinically relevant variants with a variant allele frequency (VAF) cutoff at 2%. All the samples were also screened for ASXL1 (including c.1934dupG; p.G646WfsX12) and SRSF2 mutations by Sanger sequencing. Moreover, aligned reads from .bam files were visualized using the Integrative Genomics Viewer v2.3 from the Broad Institute (Cambridge, MA, USA). Assessment of variant implication was performed based on population databases (dbSNP and GnomAD), mutation databases (COSMIC), and prediction software (Alamut, mutation taster, OncoKB, and Cancer Genome Interpreter).

Dussiau C., Boussaroque A., Gaillard M., Bravetti C., Zaroili L., Knosp C., Friedrich C., Asquier P., Willems L., Quint L., Bouscary D., Fontenay M., Espinasse T., Plesa A., Sujobert P., Gandrillon O, & Kosmider O. (2022). Hematopoietic differentiation is characterized by a transient peak of entropy at a single-cell level. BMC Biology, 20, 60.

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Genetic Profiling of Neurological Disorders

Cited 2 times

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A subset of our patient participants actively consented to donate blood and saliva to the Scottish Regenerative Neurology Tissue Bank for genetic analysis. Use of patient samples for genetic profiling has been approved by the Chief Scientist Office Scotland; MREC/98/0/56 1989–2010, 10/MRE00/77 2011–2013, 13/ES/0126 2013–2015, 15/ES/0094 2015-present. C9orf72 genotyping was performed as described in [12 (link)] and most cases were genetically analyzed as described in [5 (link)]. Some cases were analyzed by next generation sequencing to screen for variants in the coding regions of SOD1, TARDBP, FUS, VCP, UBQLN2, SQSTM1, CHMP2B and VAPB. Library construction was carried out using the Ion Ampliseq library kit v2.0 (Life Technologies). Emulsion PCR and enrichment was performed using the Ion OneTouch2 instrument (200p template kit) and Ion ES module. Sequencing was carried out on an Ion Torrent Personal Genome Machine (Life Technologies) using the Ion PGM sequencing 200 kit v2 and an Ion 316 chip. All stages followed the manufacturer’s protocols. NextGENe software (Softgenetics) was used with hg19 (GRCh37) human genome as reference to identify variants.

Henstridge C.M., Sideris D.I., Carroll E., Rotariu S., Salomonsson S., Tzioras M., McKenzie C.A., Smith C., von Arnim C.A., Ludolph A.C., Lulé D., Leighton D., Warner J., Cleary E., Newton J., Swingler R., Chandran S., Gillingwater T.H., Abrahams S, & Spires-Jones T.L. (2017). Synapse loss in the prefrontal cortex is associated with cognitive decline in amyotrophic lateral sclerosis. Acta Neuropathologica, 135(2), 213-226.

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Targeted NGS Profiling of Common Oncogenes

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DNA was extracted from formalin-fixed, paraffin embedded tumour tissue after deparaffinization in xylene and rehydration in ethanol using the commercial DNA Sample Preparation Kit (Roche, Basel, Switzerland) according to the manufacturer's protocol. Mutation analysis was performed by Massively Parallel Sequencing (NGS). Indexed Illumina NGS library was constructed from 100 ng tumour cell line DNA using KAPA Library Preparation Hyper Plus Kit (Kapa Biosystems). Hybrid selection was performed with a custom SeqCap EZ Choice Library (Roche NimbleGen). The library was designed using the genome build hg19 NCBI Build 37.1/GRCh37 (input genomic regions: KRAS NM_004985.4 -full coding regions; NRAS NM_002524.4 -full coding regions; BRAF NM_004333.4 -exons 11 a 15; PTEN NM_000314.4 -full coding regions). Paired-end cluster generation and sequencing were performed according to standard protocols from Illumina, using v2 kits. Sequencing data analysis and variant annotation was performed using NextGENe software (Softgenetics) with minimum 5% variant allele frequency filtering.

Krbal L., Soukup J., Stanislav J, & Hanusova V. (2017). Derivation and basic characterization of colorectal carcinoma primary cell lines. Biomedical papers of the Medical Faculty of the University Palacky, Olomouc, Czechoslovakia, 161(4).

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Comprehensive Genetic Profiling of Myeloid Malignancies

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Example 3

Using Sanger sequencing and NGS, mutations were analyzed in the following genes: ASXL1, ETV6, EZH2, IDH1, IDH2, NRAS, CBL, RUNX1, SF3B1, SRSF2, TET2, TP53, U2AF1, and ZRSR2. Sanger sequencing was performed using a standard protocol. The primer pairs were designed to encompass >90% of reported mutations in these genes. Polymerase chain reaction (PCR) products were purified and sequenced in both forward and reverse directions using an ABI PRISM 3730XL Genetic Analyzer (Applied Biosystems, Foster City, Calif.). Sequencing data were base-called using sequencing analysis software and assembled and analyzed with SeqScape software (Applied Biosystems).

NGS was performed using the Illumina MiSeq system (San Diego, Calif.); NGS, amplification, and indexing were performed as recommended by the manufacturer. Amplicons were confirmed for each sample by running an agarose gel. Samples were pooled and the experiment sheet was generated using Illumina Experiment Manager. MiSeq Reporter was used for analysis and Variant Studio was used for calling. For confirmation of variant calling, NextGene software (SoftGenetics, State College, Pa.) was used. Average sequencing coverage across the entire coding regions was 4,000 in 94% of the sequenced amplicons. This reliably allowed detection of mutations if present in at least 3% of mutant DNA.

US10604801B2. Deep sequecing of peripheral blood plasma DNA as a reliable test for confirming the diagnosis of myelodysplastic syndrome (2020-03-31). NeoGenomics Laboratories, Inc. [US]. Inventors: Maher Albitar [US].

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Multilevel Genomic Profiling of Glioma Samples

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DNA from glioma cells was extracted using the commercial DNA Sample Preparation Kit (Roche, Basel, Switzerland). Mutation analysis was performed by multi parallel sequencing (NGS) using the hybrid-capture-based target enrichment. A custom KAPA HyperChoice MAX Library (Roche) for enrichment of the coding and 30 bp upstream and downstream overlaps of selected panel of genes (TP53, EGFR, IDH1, IDH2, PTEN, PIK3CA) was used. Paired-end cluster generation and sequencing was performed by NGS system Illumina MiniSeq. Sequencing data analysis were performed by NextGENe software (Softgenetics) and Varsome Clinical Platform. MGMT methylation analysis was performed on DNA in FFPE using DNA Sample Preparation Kit (Roche). Bisulfite conversion of isolated DNA was performed using the EZ DNA Methylation-Gold Kit (Zymo Research). Detection and quantification of the hypermethylation status of the O (6)-methylguanine-DNA methyltransferase (MGMT) promoter was performed by the methylation-specific real-time PCR method using the CE-IVD marked geneMAP MGMT Methylation Analysis Kit (GenmarkSalgik).

Dvorakova K., Skarkova V., Vitovcova B., Soukup J., Vosmikova H., Pleskacova Z., Skarka A., Bartos M.C., Krupa P., Kasparova P., Petera J, & Rudolf E. (2024). Expression of STAT3 and hypoxia markers in long-term surviving malignant glioma patients. BMC Cancer, 24, 509.

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Targeted NGS Profiling of Myeloid Malignancies

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BCR-ABL1 KD amplicon libraries were prepared using the Nextera XT DNA Library Prep Kit (cat. number FC-131-1096, Illumina, San Diego, CA, USA).
The DNA custom-designed NGS panel of 33 genes often mutated in myeloid malignancies (Supplementary Table S2) was analyzed using SeqCap EZ HyperCap Workflow (Roche, San Diego, CA, USA).
Sequence analysis and mutation identification were performed using NextGENe software (SoftGenetics, State College, PA, USA).

Curik N., Polivkova V., Burda P., Koblihova J., Laznicka A., Kalina T., Kanderova V., Brezinova J., Ransdorfova S., Karasova D., Rejlova K., Bakardjieva M., Kuzilkova D., Kundrat D., Linhartova J., Klamova H., Salek C., Klener P., Hrusak O, & Machova Polakova K. (2021). Somatic Mutations in Oncogenes Are in Chronic Myeloid Leukemia Acquired De Novo via Deregulated Base-Excision Repair and Alternative Non-Homologous End Joining. Frontiers in Oncology, 11, 744373.

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Genetic Variant Identification in Cardiomyopathy

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Peripheral blood was collected and genomic DNA was extracted according to standard procedures. Oligonucleotide-based target capture (Agilent SureSelect Target Enrichment System; Agilent, Santa Clara, California, United States) and subsequently NGS (Illumina HiSeq2500) were used to identify potential variants of 62 genes implicated in the causation of cardiomyopathy as described previously [8 (link)]. Alignment of sequence reads to reference human genome (Human 37.3, SNP135) was performed using the NextGENe® software (SoftGenetics, Stage College, Pennsylvania, USA). All single nucleotide variants and indels were saved as VCF format files, and uploaded to Ingenuity® Variant Analysis™ (Ingenuity Systems, Redwood City, California, USA) for variations filtering and interpretation. All the variations were classified according to the recommended method of the American College of Medical Genetics and Genomics. Pathogenic and potentially pathogenic mutations were confirmed by Sanger sequencing, where possible, validated by parental testing and segregation analysis. NM_000116.3 was used as the reference sequence for the coding regions of the TAZ gene. There was an approximate 4–6 week-period from laboratory receipt to report generation.

Wang J., Guo Y., Huang M., Zhang Z., Zhu J., Liu T., Shi L., Li F., Huang H, & Fu L. (2017). Identification of TAZ mutations in pediatric patients with cardiomyopathy by targeted next-generation sequencing in a Chinese cohort. Orphanet Journal of Rare Diseases, 12, 26.

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Genetic Testing for Cardiomyopathy Disorders

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Peripheral blood was collected, and genomic DNA was extracted according to standard procedures. Genetic testing was performed by next-generation sequencing (NGS) using either an expanded cardiomyopathy panel (58 genes, including 12 known RASopathy genes: BRAF, CBL, HRAS, KRAS, MAP2K1, MAP2K2, NF1, NRAS, PTPN11, RAF1, SHOC2, and SOS1) or a whole-exome sequencing approach (Illumina HiSeq2500). Alignment of sequence reads to a reference human genome (Human 37.3, SNP135) was performed using the NextGENe® software (SoftGenetics, State College, Pennsylvania, USA). All single nucleotide variants and indels were saved as VCF format files, and uploaded to Ingenuity® Variant Analysis™ (Ingenuity Systems, Redwood City, California, USA) for variation filtering and interpretation. All the variations were classified according to the recommended method of the American College of Medical Genetics and Genomics [7 (link)]. Pathogenic and potentially pathogenic mutations were confirmed by Sanger sequencing, where possible, and validated by parental testing and segregation analysis. Variants classified as likely benign or benign were not selected to validate using Sanger sequencing.

Chen H., Li X., Liu X., Wang J., Zhang Z., Wu J., Huang M., Guo Y., Li F., Wang X, & Fu L. (2019). Clinical and mutation profile of pediatric patients with RASopathy-associated hypertrophic cardiomyopathy: results from a Chinese cohort. Orphanet Journal of Rare Diseases, 14, 29.

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