The aims of our study were 1 to compare the performance of ampliseq and sureselect and 2 to develop an optimized protocol for variant calling for wes on ion. Pdf progress in ion torrent semiconductor chip based. These amplicons are sequenced on the ion pgm, allowing for rapid turnaround and low cost. Here, we describe several examples of how these white glove initiatives have resulted in technological advances which will expand the repertoire of targeted rna sequencing applications. Ion pi sequencing 200 kit v2 kansas state university. Processing ion ampliseq data using nextgene software v2. The detailed protocol was described elsewhere usami et al. Frontiers evaluation of the performance of ampliseq and. Configurable to a specific human disease area, and ordered in small reaction packs. Ampliseq for illumina works with dna and rna samples and requires as little as 1 ng of input. The touchtools application starter screen lets the operator select the desired protocol with the touch of a button.
Thermo fisher scientific and illumina sign agreement to. For rnaseq libraries, the quality of input rna, purification. We adapted our ion rna ampliseq primer designer to enable design to multiple. Ion ampliseq dna custom panels enable affordable targeted sequencing for your genomic region of interest. With emphasis on validation with relevant single molecular pathology tests. Get the benefit of a comprehensive, growing menu of content, scalable sequencing platforms and local or cloudbased analysis solutions. Illumina ilmn ties up with thermo fisher for ion ampliseq.
The ion ampliseq cancer hotspot panel v2 is now expanded to allow translational and disease researchers to fasttrack oncology research by surveying hotspot regions of 50 oncogenes and tumor suppressor genes, with wide coverage of the kras, braf, and egfr genes. Ampliseq for illumina comprehensive cancer panel, store at 25c to 15c. Ampliseq panels can accommodate highquality samples such as blood, cell culture, or fresh frozen tissue and also challenging samples such as formalinfixed paraffinembedded ffpe tissue. A free design tool that helps you obtain custom assay designs based on pcr target selection ion ampliseq designer. Clinical application of a custom ampliseq library and ion. 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. Thermo fisher will continue to sell ion ampliseq chemistry for both ivd and ruo applications to ion torrent ngs customers, and retains the right to make the technology available on other nextgeneration sequencing platforms. The ion ampliseq library kit plus is designed for rapid preparation of amplicon libraries using ion ampliseq panels. Communications protocol and ion modbus register map this document explains the modbus protocol on the ion6200 meter. With low dna input, the panel provides high accuracy, ease of use, and short time to answer.
The performance of this panel has previously been evaluated for clinical cancer testing by other groups retrospectively on archival16 and clinical trial specimens. This panel, with improved primer design, contains 207 primer pairs in a single tube and. Determining genotypic drug resistance by ion semiconductor. The protocol is simple to use and enables high yield and uniformity, reliably and reproducibly. Looking through and comparing the steps of the protocol, ion ampliseq has 59 protocol steps, while pillars slimamp has only 34.
Each of these reagents has been functionally validated by preparation of a genomic dna library followed by ion torrent sequencing. Ion ampliseq dna and rna library preparation user guide pub. Ampliseq designer and any custom references you have uploaded. For representative results of both library construction using nebnext and ion torrent run results, choose the tabs below. The ampliseq for illumina comprehensive cancer panel includes the following reagents. You can access this calculator either by the navigation bar at the top of the screen, or on individual panel pages. Process extracted dna, rna or ffpe samples through library prep and pooling with the ion torrent ion chef system or manual preparation. Ion ampliseq technology enables scalable, multiplex polymerase chain reactions pcr from 12 to 24,000plex in a single well.
Designed for individuals preparing ion ampliseq libraries for sequencing on the ion torrent systems. Evaluation of the performance of ampliseq and sureselect exome. Ion ampliseq comprehensive cancer panel facilitates broad oncology surveys using only 40 ng of dna with 16,000 primer pairs multiplexed into just 4 pools. This can reduce the amount of filtering that is needed and increase the percentage of. This guide contains everything a firsttime user needs, including an overview of the ampliseq for illumina exome panel protocol, tips and techniques, protocol steps, kit contents, and usersupplied consumables. 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. In the select a report format section, select the pdf or text. Ion ampliseq rna cancer panel was developed as an rna complement to the ion ampliseq cancer hotspot panel v2. Ion ampliseq direct ffpe dna kit ion torrent dual barcode kit 196 ioncode barcode adapters 84 kit ion xpress barcode adapters kits ion library equalizer kit ion ampliseq readytouse panels ion ampliseq madetoorder and community panels. Ion proton system, ion pgm system, and ion s5 system. Man0006735 for detailed instructions, including protocols for 3. Whole transcriptome sequencing rnaseq represents a powerful approach for whole transcriptome gene expression analysis. Briefly, after amplifying the target region using 12 pools of ion ampliseq primers 24,000 primer pairs totaling to 294,000, we partially digested. Amplify target regions from dna or cdna with the ion ampliseq library kit and ion ampliseq readytouse, custom, or community panel.
This library kit is compatible with the following instruments. How does ampliseq ngs enrichment compare to pillar. The ampliseq for illumina solution offers a highly multiplexed polymerase chain reaction pcrbased workflow for use with targets ranging from a few to hundreds in a single run. Panels can be designed using a number of preloaded reference genomes, or you can upload your own reference sequence for any other organism. When using the ion torrent pairedend protocol, the overlapping reads can be merged into highquality single reads in order to improve sequence accuracy. Use of the ion ampliseq cancer hotspot panel in clinical molecular pathology laboratories for analysis of solid tumours. The overall illumina ion ampliseq workflow is available as a handy graphic see this reference guide pdf online. Analysis of ampliseq rnasequencing enrichment panels. Aug 11, 2015 hear scott dewell, product manager for rna applications with the ion torrent group, describe how easy it is to analyze nextgen sequencing data from the ion ampliseq transcriptome solution.
Apr 07, 2015 one of the advantages of ion ampliseq library preparation is thought to be this high assay success rate. Pdf use of the ion ampliseq cancer hotspot panel in. The ion torrent dx cdna synthesis kit and the oncomine dx target test and controls are used to reverse transcribe the quantified rna into cdna. On demand ion ampliseq research panels of optimized amplicons for germline analysis. The panel is a single pool of primers targeting 50 oncogenes and tumor suppressor genes including the kras, braf, and egfr genes. Aug 07, 2018 looking through and comparing the steps of the protocol, ion ampliseq has 59 protocol steps, while pillars slimamp has only 34. Added support for the ion ampliseq direct ffpe dna kit. Ion ampliseq targeted sequencing technology thermo fisher. Madetoorder ion ampliseq research panels of dna and rna designs for germline or somatic analysis of.
Thus, the aim of our study was to evaluate the efficiency of a fast protocol based on custom ampliseq panel and ion pgm sequencer for sequencing most prevalent cmcausing genes myh7, mybpc3, myl2, lmna, scn5a, tnnt2, tnni3 and tpm1. Pdf in order for nextgeneration sequencing to become widely used as a. This chapter describes some procedures and tools used in ion ampliseq designer. Ion ampliseq rna cancer panel targets 50 oncogenes with detection of key genes kras, braf, and egfr.
Easy and robust protocol the ion ampliseq library kit plus uses a platebased format for easier sample handling and tracking, and for compatibility with automation and highthroughput laboratories. Thermo fisher will continue to sell ion ampliseq chemistry for both ivd and ruo applications to ion torrent. Automation protocol for information about preparing ion ampliseq libraries on the tecan freedom. Jun 10, 2014 thus, the aim of our study was to evaluate the efficiency of a fast protocol based on custom ampliseq panel and ion pgm sequencer for sequencing most prevalent cmcausing genes myh7, mybpc3, myl2, lmna, scn5a, tnnt2, tnni3 and tpm1. A cohort of previously studied 75 cm patients was screened in order to evaluate this strategy in terms of. Ampliseq for illumina comprehensive cancer panel contents. Ion ampliseq dna custom panels thermo fisher scientific. If preparing ion ampliseq exome libraries, see the protocols listed on page 7. Studies comparing the only two library preparation methods, namely, ampliseq and sureselect for ion proton, are, however, so far missing.
The ion6200 meter performs modbus rtu communications according to the modicon standard. Encompassing over 50% of the wellcome trust sanger institute cancer gene census, this is the most comprehensive cancer gene panel available. 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. These library kits use a platebased protocol for easy sample handling, and for compatibility with automation in high throughput. Processing ion ampliseq cancer panel data using nextgene. Ion torrent sequencing microarray and sequencing resource. Ampliseq for illumina is a targeted resequencing solution that delivers highly accurate data for a wide range of sample types, including ffpe tissue and low sample input. Comprehensive evaluation of ampliseq transcriptome, a. Change product name to hidion ampliseq library kit.
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. Ion ampliseq library kit plus thermo fisher scientific. Scalable multiplex pcr reactions the ion ampliseq 2. Ion ampliseq rna cancer panel thermo fisher scientific. Portable document format pdf versions of this guide and the documents listed above.
The ion ampliseq library preparation used in this study required only 20 ng dna samples, and the quality of the dna samples did not. 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. Jan 15, 2018 illumina ilmn ties up with thermo fisher for ion ampliseq. 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. Easy analysis of ion ampliseq transcriptome data youtube. Ampliseq for illumina is a suite of ampliseq chemistry products that are compatible with illumina nextgeneration sequencing platforms. One way this can be illustrated is with looking at the reagents each sequencing enrichment kit contains. Ion ampliseq dna and rna library preparation user guide pn. Ion ampliseq dna and rna library preparation user guide for use with. Simply spike any pcr multiplex reaction with 1 l of the panel and continue with the standard ion ampliseq protocol. Evaluation of the performance of ampliseq and sureselect.
Ion ampliseq hd library kit thermo fisher scientific. Sample preparation and quantification oncomine dx target test. 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. The ion ampliseq transcriptome human gene expression kit ampliseq was recently. Ion ampliseq dna and rna library preparation user guide 7. To assess the performance of this new methodology, we. Ion ampliseq library kit 2 thermo fisher scientific. The aims of our study were 1 to compare the performance of ampliseq and sureselect and 2 to develop an optimized protocol for variant calling for wes on ion proton platform. A free design tool that helps you obtain custom assay designs based on pcr target selection. The existence of potentially significant variants was further reassessed through manual inspection of aligned reads in igv 2. Comprehensive evaluation of ampliseq transcriptome, a novel. See ampliseq for illumina comprehensive cancer panel compatible products for information on compatible products, including index and library prep kits. Lets find out what this new technology can do for you.
Nextgeneration sequencing ngs libraries can be prepar. The ion ampliseq transcriptome human gene expression kit ampliseq was recently introduced by life technologies as a wholetranscriptome, targeted gene quantification kit to overcome these limitations of rnaseq. One of the advantages of ion ampliseq library preparation is thought to be this high assay success rate. Clinical application of a custom ampliseq library and ion torrent pgm sequencing to comprehensive mutation screening for deafness genes. The new ion ampliseq exome kit delivers the best exome sequencing performance in 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. Manual inspection of all fns on chr 1, 7, 16, 19, and x. Mar 05, 2020 to our knowledge, we were the first to adapt sureselect hybridisationbased enrichment protocol for the sequencing on the ion torrent s5 platform, which is intended to work preferably with ampliconbased panels f. Ion ampliseq library preparation on the ion chef system user.
Ion torrent is a trademark owned bythermo fischer scientific. Hear scott dewell, product manager for rna applications with the ion torrent group, describe how easy it is to analyze nextgen sequencing data from the ion ampliseq. Ion reporter ampliseq multiplex pcr ion ampliseq sample id panel simple and accurate sample identification for downstream verification figure 1. 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. Attendees will benefit by having training in a class environment, handson lab practicals, and interaction with fellow attendees and ion torrent application scientists during lecture, discussion, and lab sessions. Manual library preparation using the ion ampliseq library kit plus cat. The ngs library was constructed using the ion ampliseq library kit 2.
Lowdna input protocol and a singleday ion ampliseq workflow. Five independent janus protocols are used in the ion ampliseq library preparation workflow. Working with key cancer researchers and incorporating information from literature. Thermo fisher scientific launches ion ampliseq hd technology. Access codes are used to integrate between all ion torrent software. Adapting sureselect enrichment protocol to the ion torrent s5.
Sample preparation and quantification oncomine dx target. Andy felton, ion torrents head of product marketing, explains how the new ion ampliseq exome kit works. The ion ampliseq library prep application manages sample data for the ion ampliseq library and ion ampliseq hd library preparation workflows. Progress in ion torrent semiconductor chip based sequencing. Ion ampliseq designer home page before you start a panel design or browse predesigned panels, sign in at ampliseq. Additional dilution points may be required for accurate quantification if alternate protocols are employed. Highly multiplexed ampliseq technology identifies novel variation of. Get started with ion ampliseq designer welcome to ion ampliseq designer. Library preparation was done with the ion ampliseq library kit plus life technologies, usa using 100 ng of dna 100 ngl according to the manufacturers protocol. Evaluation of a new ngs method based on a custom ampliseq. Ion ampliseq dna and rna library preparation user guide. Illumina will sell the product directly to its customers under the name ampliseq for illumina. Ion ampliseq library preparation for human identification applications.
Ion torrent semiconductor sequencing is performed on either the personal genome machine pgm or proton instrument depending on desired read length and application. Getting started user guide 7 you can also browse predesigned community panels in the same research areas by clicking the predesigned by our community button in the madetoorder. This ion ampliseq methylation panel protocol combines a one day bisulfite kit protocol with the one day ion ampliseq library kit plus protocol to create ngs libraries for target dna methylation assessment during sequencing. Library preparation was done with the ion ampliseq library kit plus life. Ion ampliseq sample id panel thermo fisher scientific. Novel cutting edge applications of ion ampliseq rna. Thermo fisher will continue to sell ion ampliseq chemistry for both ivd and ruo applications to ion. Ion ampliseq ion ampliseq panels how is a protocol started. Ion ampliseq library prep platform for science marketplace. The ion ampliseq transcriptome human gene expression kit ampliseq was recently introduced by life. Purpose of the guide the ion pi sequencing 200 kit v2 user guide pub. Following protocol selection, specific run information including any variable number of. Prepare dna with the ion ampliseq direct ffpe dna kit. Processing ion ampliseq cancer panel data using nextgene software introduction the ion ampliseq cancer panel uses highly multiplexed pcr in order to generate amplicons from 46 different cancer genes.
1462 127 20 1265 1412 1303 499 1098 245 792 149 1498 833 823 580 569 235 1181 849 1303 560 466 1203 344 776 701 1072 1491 832 442 674 1257 946 173 1078