As next generation sequencing evolves, the ability to create reproducible, high quality libraries becomes increasingly important. Preexisting or customized primer pools allow highly multiplexed pcr amplification of thousands of genomic target regions, with superior coverage uniformity and specificity. Evaluation of the performance of ampliseq and sureselect. Ampliseq for illumina library prep, index adapters, and. Ion ampliseq library prep comes to the ion chef agbt. Thermo fisher scientific launches ion ampliseq hd technology. For information about preparing ion ampliseq libraries on the tecan freedom. Many researchers have the perception that nextgeneration sequencing systems are complex and that managing the workflow and analysis requires extensive training. Component amount storage 5x ion ampliseq hifi mix red cap 120 l 30. Automated, purificationfree, ion ampliseq library preparation from ffpe samples using the ion torrent ion chef system. The initial input requirement is the same as the ion ampliseq protocol. And i should mention too, after library preparation, performing an additional library quantification is key. Library preparation is a critical, handson and timeconsuming step in the ngs workflow. Lets find out what this new technology can do for you.
Each of these reagents has been functionally validated by preparation of a genomic dna library followed by ion torrent sequencing. Thermo fisher scientific and illumina sign agreement to. The ampliseq for illumina library plus kit is available in 24, 96, and 384reaction configurations. Jun 19, 2015 as next generation sequencing evolves, the ability to create reproducible, high quality libraries becomes increasingly important.
These library kits use a platebased protocol for easy sample handling, and for compatibility with automation in high. The panel, created using the ion ampliseq designer software, was designed to identify diseasecausing mutations in the following genes. Ion training courses cover library preparation, template preparation, sequence generation, and data analysis and are offered at the training centers in carlsbad, california and frederick. Process extracted dna, rna or ffpe samples through library prep and pooling with the ion torrent ion chef system or manual preparation. During library preparation, each library is prepared in an independent well of a 96well plate, encompassing several washes and magnetic beadbinding steps. These terms and conditions supplemental primer terms apply to your use of products and materials provided to you by life technologies corporation hereinafter, ltc and that include any of the following. One of the advantages of ion ampliseq library preparation is thought to be this high assay success rate. Ion ampliseq dna and rna library preparation user guide pn. There are several important differences between the ion ampliseq protocol and the ampliseq for illumina protocol, such as added amplification and cleanup steps. Nextgeneration sequencing ngs libraries can be prepar. Course covers handson lab training of ultrahigh multiplex pcr of targets using ion ampliseq technology, adapter ligation, amplification, and quantification methods.
The ion ampliseq library prep application manages sample data for the ion ampliseq library and ion ampliseq hd library preparation workflows. This difference adds approximately 4560 minutes total time to ampliseq for illumina library prep included in above estimates. Amplicon sequencing is the primary supported dna sequencing application for the accelngs dna library kit for ion torrent. Application guide ion ampliseq library preparation on the ngs express perkinelmer 5 because there are several points at which the samples can be stored in the process, the overall library preparation was broken down into multiple smaller steps. Ion ampliseq library preparation on the ion chef system user guide. Ampliseq for illumina exome panel support resources. Evaluation of a new ngs method based on a custom ampliseq. Ion proton system, ion pgm system, and ion s5 system.
Fully automated ion ampliseq library preparation using the. Amplify target regions from dna or cdna with the ion ampliseq library kit and ion ampliseq readytouse, custom, or community panel. Although of major interest, a comparison of the two methods is hitherto missing in the literature. Ion ampliseq library kits ion ampliseq library kit 2. Ampliseq for illumina cancer hotspot panel v2 50 hotspot. Ion ampliseq library preparation on the ion chef system. Apr 01, 2015 one of the advantages of ion ampliseq library preparation is thought to be this high assay success rate. Allen, nathan kane, darren birr, tanya biorac, ryan jones, maryam shenasa, edward liu, sylvia do, david marran, kate rhodes, and mark andersen life sciences solutions, thermo fisher scientific fully automated ion ampliseq library preparation using the ion chef system introduction. Ion ampliseq library preparation for human identification applications user guide 5 about this guide important.
The ion ampliseq library preparation used in this study required only 20 ng dna samples, and the quality of the dna samples did not affect the sequence results. The answer is a next generation sequencing, or ngs, library. The accelngs dna library kit designed for the ion torrent platform is the only commerciallyavailable kit capable of producing pcrfree libraries with as little as 5 ng of input dna. Frontiers evaluation of the performance of ampliseq and. Ion ampliseq library preparation on the ion chef system catalog numbers a29024, a35121, 4484177, a32914, a29998, a31446, a32841 pub. About thermo fisher scientific thermo fisher scientific inc. Products by brand ampliseq for illumina community panels ampliseq for illumina inherited cancer research panel ampliseq for illumina inherited cancer research panel this sequencing research panel provides a quick, accurate, and costeffective way to identify genetic variants in 4 genes associated with inherited cancers. Miniaturization technologies for costeffective ampliseq library. The ion ampliseq library kit plus is designed for rapid preparation of amplicon libraries using ion ampliseq panels. Ion pgm has accessory equipment that might no longer be needed.
Ion ampliseq technology enables scalable, multiplex pcr reactions from 12 to 3,072plex in a single tube using just 10 ng of starting dna. Ion ampliseq library preparation simplified eshg 2015 youtube. Before using this product, read and understand the information the. From predesigned to customizable dna and rna panels for nextgeneration sequencing ngs, all of our content is available within the ion ampliseq designer including indepth descriptions and gene lists for each panel. Background morphologic and genetic analysis of thyroid nodules may be performed from a single vial. Ion ampliseq library preparation is available in ion ampliseq library kit 2.
Prepare an ontarget, highuniformity amplified library. The ion ampliseq method has proven useful to reliably generate. Ampliseq for illumina library prep kit definition files for local run manager. Sep 25, 2019 library preparation for wholeexome sequencing is a critical step serving the enrichment of the regions of interest. Evo liquid handling platform, see prepare ion ampliseq. Libraries are then ready for template preparation on the ion onetouch system and sequencing on the ion pgm sequencer. The myeloid panel is part of an integrated workflow that includes ampliseq for illumina polymerase chain reaction pcrbased library preparation, illumina sequencing by synthesis sbs, ngs technology, and automated analysis. With the ion s5 sequencers, ion chef system, and ion ampliseq technology, you get the simplest targeted sequencing workflow. Library preparation for wholeexome sequencing is a critical step serving the enrichment of the regions of interest. Instructions for using the ampliseq for illumina cancer hotspot panel v2. A38262 and a38264 include the ion ampliseq library kit plus and the ion ampliseq exome rdy panel, which provide reagents for preparing eight exome libraries. User guide ion ampliseq library preparation is available in ion ampliseq library kit 2. Jun 25, 2015 fully automated ion ampliseq library preparation using the ion chef system 1.
Automated library preparation of ion ampliseq panels many researchers have the perception that nextgeneration sequencing systems are complex and that managing the workflow and analysis requires extensive training. Introduction to sample preparation and library qc for ion. Next generation sequencing is now producing more sequence, faster than anyone. Automated library preparation of ion ampliseq panels. Ion ampliseq exome rdy kit 4x2 thermo fisher scientific. For copy number analysis of ion ampliseq data, ion reporter software thermofisher scientific, ma, usa is widely used. For dna library preparation, reagents from neb are available with or without reagents for enzymebased fragmentation. Ion ampliseq library preparation quick reference 3 quantify the library and dilute determine the concentration of each ion ampliseq library by qpcr with the ion library quantitation kit cat. After ligation, adaptordna constructs are purified and size selected, and amplified via polymerase chain reaction pcr. Demand panels ion ampliseq readytouse panels ion ampliseq madetoorder and community panels. Fast library prep, optimized for small genomes, pcr amplicons, and plasmids. The ion ampliseq kit for chef dl8 dna to library, 8 samplesrun includes the reagents and materials for automated preparation of up to 8 barcoded ion ampliseq libraries per ion chef run using a single 1 or 2pool ion ampliseq panel. Detect somatic mutations down to 5% frequency using local or cloudbased analysis. Users who are interested in inherited disease or germline research applications can design and order panels using.
Ion ampliseq dna and rna library preparation user guide chapter 1 product information procedure overview 1 procedure overview if starting with rna, first reversetranscribe 10 ng to make cdna. Preexisting or customized primer pools allow highly multiplexed pcr amplification of thousands of genomic target regions, with superior coverage uniformity and. This has led thermo fisher to develop ion ampliseq hd, a new library prep technology that enables custom design flexibility for single nucleotide variations snvs, small insertionsdeletions indels, fusions and copy number variation cnv detection with a limit of detection lod as low as. Ion ampliseq exome rdy kits the ion ampliseq exome rdy kits cat. This oneday course features classroom and laboratory sections for ion ampliseq library construction. Application guide ion ampliseq library preparation on the ngs express perkinelmer 4 introduction the janus ngs express ion ampliseq application is designed for automated preparation of amplicon libraries using ion ampliseq panels and custom primer pools and then sequencing on the ion personal genome machine pgm system.
Demand panels are designed for use with the ion ampliseq library kit plus for manual dna library preparation, or the ion ampliseq kit for chef dl8 for automated dna library preparation. Ion ampliseq library preparation on the ion chef system user guide for use with. Validating ngs library preparation with ion ampliseq kit. Ngs library was constructed using the ion ampliseq library kit 2. Jun 10, 2014 library preparation genomic dnas were tested by ngs sequencing using a custom design for the cardiomyopathy panel, based on an ampliseq strategy lifetechnologies, carlsbad, ca, usa. Ion ampliseq technology enables scalable, multiplex polymerase chain reactions pcr from 12 to 24,000plex in a single well. Dna library preparation with 1 or 2pool panels using qpcr quantification. In addition, ion ampliseq uses multiplex pcr products as sequencing libraries, and most of the target sequence is covered by only one amplicon. Nextgeneration sequencing ngs libraries can be prepared using as little as 1 ng of starting material.
Illumina benchtop sequencing consumables are single cartridgebased with a separate flow cell. The equipment needed for performing an ampliseq for illumina library prep is the same as the equipment needed for performing an ampliseq library experiment on the ion torrent. The flexible ion chef system is designed to provide automated and consistent ion ampliseq library preparation and template preparation for sequencing on the ion genestudio s5 systems using any of the corresponding chips or template preparation and sequencing chemistries. Scalable multiplex pcr reactions the ion ampliseq 2. Ampliseq for illumina cancer hotspot panel v2 documentation. Five independent janus protocols are used in the ion ampliseq library preparation workflow. Ion ampliseq kit for chef dl8 ion ampliseq chefready kits oncomine research assay chefready kits ion ampliseq direct ffpe dna kit ion ampliseq on. Manifest files for the ampliseq for illumina cancer hotspot panel v2. This kit uses only certain components found in the ion ampliseq library kit 2. Ampliseq for illumina inherited cancer research panel.
The ion ampliseq exome rdy kits contain reagents for the rapid preparation of eight exome libraries for sequencing using the one of the ion genestudio s5 series sequencers, the ion s5 ion s5 xl sequencer, or the ion proton sequencer. For ion proton, there are only two exome library preparation methods available, ampliseq and sureselect. Ion ampliseq library preparation for human identification. Trusight, and ampliseq for illumina library prep kits. The latest addition to the illumina library prep portfolio, the ampliseq for illumina solution provides a simple, fast, and robust amplicon sequencing option to achieve highconfidence data from. Demonstration of library prep protocol steps and best practices. The illumina protocol has a second 7cycle pcr step at the end of the protocol and an additional round of bead cleanup when compared to the thermo fisher ion ampliseq protocol. Ion ampliseq library preparation on the ion chef system user. In general, library preparation of fragmented dna, for use with the ion torrent sequencing platform, involves repair of 3. Transitioning from ion torrent to illumina ngs platforms. Thermo fisher scientific launches ion ampliseq hd technology new library preparation solution for targeted nextgeneration sequencing is first to combine gene panel design customization with ultra. Tmo launches its ion ampliseq hd technology, the latest nextgeneration library preparation that, provides the flexibility to custom design gene panels without sacrific.
Library preparation with ampliseq for illumina is fast and simple. A multiplexed, polymerase chain reaction pcrbased workflow replaces nonspecific hybridization steps, resulting in a highspecificity, highuniformity amplified library. For information on index kit compatibility, refer to the reference guide for your panel. Clinical application of a custom ampliseq library and ion. Ampliseq for illumina library plus and indexes are optimized for use with illumina sequencers, and generate highly accurate data with an extensive menu of readytouse or made to order content. Proceedings of the 107th annual meeting of the american association for cancer research. Ampliseq for illumina cancer hotspot panel v2 product files.
Dna library preparation with 1 or 2pool panels using qubit fluorometer or agilent 2100. Prepare an ontarget, highuniformity amplified library in as little as 5 hours with just 1. Take advantage of low input from as little as 1 ng and flexible input ranges from 1 to 100 ng. Ion ampliseq library prep comes to the ion chef agbt 2015. Ion training courses cover library preparation, template preparation, sequence generation, and data analysis and are offered at the training centers in carlsbad, california and frederick, maryland. Ion ampliseq library kit plus the ion ampliseq library kit plus cat. Ampliseq chemistry allows researchers to analyze hundreds of genes simultaneously with ultrahigh multiplexed pcr with 12. Differences between ion ampliseq and ampliseq for illumina library prep protocols. Today, we are going to focus on the four basic steps of ngs library preparation that can be broadly applied across different.
The ion torrent systems combine semiconductor sequencing technology with natural biochemistry to directly translate chemical information into digital data. From predesigned to customizable dna and rna panels for nextgeneration sequencing ngs, all of our content is available within the ion ampliseq designer including indepth descriptions and gene lists for. Preanalytic variables that affect nucleic acid extracted from a single vial are evaluated. Leveraging the ultrahigh multiplex pcr approach of ion ampliseq technology with template preparation on the ion chef system and sequencing on the ion proton or ion genestudio s5 nextgeneration sequencers, the ion ampliseq exome rdy kit allows for rapid sequencing of.
Ion ampliseq dna and rna library preparation user guide. Ion ampliseq library kit plus thermo fisher scientific. Custom training can be arranged at customer sites, and webbased instruction is also available. This is important because the ratio of dna to ion sphere particles or beads for the template preparation will dictate whether the emulsion pcr is clonal. For representative results of both library construction using nebnext and ion torrent run results. Ion ampliseq ngs panels transform genetic research with ultrahighmultiplex pcr primer pools designed for fast and simple production of thousands of targeted amplicons from samples containing as little as 1 ng of input dna, such as formalinfixed, paraffinembedded ffpe and fine needle biopsy fnb research samples. Local run manager library prep kit definition file for ampliseq for illumina panels. Next generation sequencing library preparation seq it.
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