Product Bulletin SeqScape® Software
SeqScape® Software v2.6 Detecting Mutation and SNPs, Sequence Typing
• Comprehensive
reference sequence
handling—Helps interpret the role of each polymorphism • Accurate
data analysis—Detects heterozygous indels and provides quality values
Introduction The goals of today’s large resequencing studies are to detect SNPs, profile mutations, perform medical sequencing, identify haplotypes, subtype pathogens, and confirm clone constructs. For all of
these applications, SeqScape® Software provides two levels of analysis—SNP detection and the identification of sequences that most closely match a target sequence.
• Two-tier analysis—Reduces analysis time
and complexity; identifies best sequence matches from a sequence library • Assists
with 21 CFR Part 11 compliance
NCBI graphic display of HFE Gene
—Provides data security, an audit trail of data changes, and electronic signatures • Detailed
reports—Generates comprehensive reports with detailed results describing data quality with Data Collection Software to analyze raw data generated from Applied Biosystems genetic analyzers
CDS information automatically translated in the SeqScape Reference Data Group (RDG)
• Automation—Integrated
Configure Individual Layer as transcripts Define Layer properties; Orientation, Translation Modify reference numbering
Figure 1. GenBank format file downloaded from the NCBI public database. The sequence information from the GenBank file can be directly imported into the Reference Data Group (RDG) to use as a reference sequence.
trimming, assembly, consensus alignment, and reference comparison. Character Dots View
Figure 2. For library searching, a library of pre-aligned sequences is imported into the Library Editor and aligned with the Reference Data Group (RDG). After analysis is complete, the results are reported in the Library Search Report. In the Character/Dots view of the report, base differences in the consensus are displayed against the reference as characters, and all bases common to the reference are represented as dots.
Consensus Sequence
True Variant find on bidirectional coverage
Forward Strand
Reverse Strand
Algorithm corrects the false positives and makes an edit based on bidirectional coverage
Figure 3. Example of an accurate consensus generated using base quality information from the forward and reverse strands. The consensus calling algorithm corrects for miscalls in individual strands.
Unlike tools initially designed for de novo analysis, SeqScape® Software is a comprehensive resequencing tool that analyzes data using full sequencefeature information such as exon/intron boundaries, translation frames, coding regions, and alternative transcripts. Whether your research spans multiple genes or focuses on a single region of interest, SeqScape® Software supplies you with comprehensive referencesequence features, accurate polymorphism detection, powerful viewing features, and detailed reports that can be exported for use in other analyses. Flexibility to Configure a Reference The level of relevant information to be extracted from resequencing data depends not only on the quality of subject-sample sequences, but also on the reference sequence. The Reference Data Group (RDG), an advanced feature in SeqScape Software, offers powerful capabilities for configuring reference sequence. For instance, you
can directly import a GenBank file as a reference (Figure 1). The software will automatically parse feature information from the file, including exon and intron boundaries, and translation information. This powerful feature allows the software to report mutations, nucleotides, or amino acids in the context of the genomic annotation. When studying a large gene, you can easily split the reference into smaller segments and delete any regions not required for a faster and easier data review. You can also configure alternate gene transcripts as layers with different translation frames (Figure 1). You can even create a single template to use a reference data group for multiple studies, and save time by avoiding repetitive steps.
Complete Workflow The software lets you create a new project simply by selecting a previously configured project template and adding the subject sample sequences. A click on the Analysis button is all that is required to initiate automated basecalling,
Analysis algorithms are designed and calibrated with Applied Biosystems instrument specifications and reagent characteristics to provide accurate bases with quality values for each mutation. Distinctions between poor- and highquality data are clearly defined, as the KB™ Basecaller and SeqScape Software algorithms filter out low-quality and anomalous data to provide a high-quality assembly. Quality values allow you to review your data. Using software that provides a complete workflow eliminates working with multiple tools to get to your results. Identifies Best Sequence Matches from a Sequence Library SeqScape Software compares each consensus sequence to a library of sequences and identifies the best matches, an important step in identification studies. Most labs must invest in additional software to perform this level of analysis. SeqScape® Software allows you to compare your sample data with a library of allelic sequences and perform a haploid or diploid search to identify the best matches. Software results of alignment and identification are reported within the Library Search report (Figure 2). Detects Mutations with Confidence SeqScape® Software detects all variations between the reference sequence and the subject sample sequence, including deletions, insertions, mismatches, heterozygous bases, and heterozygous insertion/deletions. The consensus caller in SeqScape® Software examines the quality of each forward and reverse trace and forms an accurate consensus call. Variations between the consensus and the reference sequence are reported as mutations in the Mutations report. Analysis at the consensus level corrects sequencing anomalies such as PCR noise or unincorporated dye terminators that are present in one strand and cause a basecalling error. Algorithms at the consensus level are essential to reduce the false-positive rate for accurate
• Displays
name when user is logged into the system
• Provides
unique user ID and password for each user
• Specifies
password lifetime
• Prevents
unauthorized user ID or password
Audit Trail Features • Generates an audit trail when bases are changed, added, or deleted Figure 4. A heterozygous deletion mutation is correctly identified and reported by SeqScape ® Software algorithms.
• Contains
user name, time and date of each change, and the reason for each change
Electronic Signatures • Captures the printed name of the signer, the date and time of the signature, and the meaning associated with the signature • Provides
a unique electronic signature for each user
Report provides hyperlinks to data in project view
Figure 5. Base changes reported in the Mutations report can be directly linked to view the sample data.
detection of heterozygote calls (Figure 3). This ensures accurate detection of all types of mutations, including heterozygous insertion or deletion mutations (HIMs), which are often missed by other methods (Figure 4). Automation Features Increase Productivity Fully integrated with Applied Biosystems Data Collection Software and reagents, which are designed expressly for our DNA sequencing instruments, SeqScape® Software, together with Data Collection Software, automates data analysis, so you can concentrate on data review. By creating a SeqScape Software plate in Data Collection Software, you can
run an automated analysis pipeline that will generate project analysis reports in SeqScape Software. Assists with 21 CFR Part 11 Requirements For researchers working in regulated environments, SeqScape® Software offers a full suite of features to provide data security, an audit trail of data changes, and electronic signatures. These features, which are designed to assist customers with 21 CFR Part 11 requirements and can be disabled or enabled at the time of software installation, include the following: Security Features • Requires user ID and password to enter the software
Detailed Reports Results reports from SeqScape® Software include hyperlinks to the data source to show the relationship between the results and the data (Figure 5). For each mutation detected, you can review the quality value provided, the classification of novel or known SNP, and the impact of the SNP on the amino acid translation level. Mutations can be sorted by each of these attributes for easy and fast review. You can also view the audit history of changes made to base pairs in the Audit report. Export Capabilities for Easy Integration SeqScape® Software provides a complete analysis, which usually requires several analysis tools. It also enables researchers to easily import results into other software packages or archival mechanisms. SeqScape Software stores results in detailed reports and enables flexible exporting options. A variety of export functionalities are provided, including alignment of the complete project, assembly across a reference segment, and individual consensus, in standard FASTA format. You can export the reports in convenient formats, such as tabdelimited text, html, and xml for easy integration into downstream pipelines.
ORDERING INFORMATION Initial License
P/N
SeqScape ® Software Initial license (Standalone or purchased with Applied Biosystems 3130/3130xl or 310 Genetic Analyzers)
4327091
SeqScape Software (3 licenses) when purchased with Applied Biosystems 3730/3730xl DNA Analyzers
4339552
®
Bundle and Save When purchased along with an Applied Biosystems capillary electrophoresis instrument, Sequencing Analysis Software can be bundled cost-effectively with our other software packages in the following combinations: 1. SeqScape® Software and Sequencing Analysis Software 2. SeqScape® Software, Sequencing Analysis Software, and GeneMapper® Software 3. SeqScape® Software, Sequencing Analysis Software, and GeneMapper® ID Software 4. SeqScape® Software, Sequencing Analysis Software, and MicroSeq ID® Analysis Software 5. SeqScape® Software, Sequencing Analysis Software, GenMapper Software and MicroSeq ID Analysis Software Call your Applied Biosystems sales representative about which combination makes the most sense for your research focus. (Terms and conditions may apply.) Related Products Variant ReporterTM Software An intuitive resequencing software to accurately discover and validate mutations and SNPs. Variant ReporterTM Software provides a task-based guided workflow and analyzes large volume data faster than other software. If your task does not require sequence library search and 21 CFR Part 11 support, Variant ReporterTM Software is a faster and more cost-effective solution than SeqScape® Software. BigDye® Terminator Cycle Sequencing Kit A robust, highly flexible chemistry, designed for the majority of applications, including de novo sequencing and resequencing. It generates data with uniform peak heights and optimized signal balance to produce long, high-quality reads. Improved peak patterns also contribute to more accurate base assignments for heterozygote and mutation detection. BigDye® XTerminator™ Purification Kit A fast, simple purification method for DNA sequencing reactions that removes unincorporated BigDye® Terminators. No more dye blobs! Clean-up is complete in under 40 minutes and requires less than 10 minutes of labor. VariantSEQr™ Resequencing System A system providing everything you need to make resequencing easy and cost effective: pre-designed PCR primer sets, universal protocols for PCR and PCR clean-up, sequencing and sequencing clean-up, and reference sequences included in the Project Template for data analysis with SeqScape® Software v2.5 or greater. Free 45-day Download and Webinars SeqScape® Software is available as a 45-day free demo for use with your own data. For more information, contact your Applied Biosystems sales representative. Free Webinars are also available to provide introductory training on the software. For a schedule of upcoming Webinars, visit: info.appliedbiosystems.com/webinars Free demo download from http://www.appliedbiosystems.com/support/software/ Reference C. Kosman et al., 2001. Design and Performance Overview of SeqScape® Software for Comparative Sequencing Analysis and Mutation Detection http://www3.appliedbiosystems.com/cms/groups/mcb_marketing/documents/generaldocuments/cms_039750.pdf
For Research Use Only. Not for use in diagnostic procedures. Notice to Purchaser: License Disclaimer Purchase of this software product alone does not imply any license under any process, instrument or other apparatus, system, composition, reagent or kit rights under patent claims owned or otherwise controlled by Applera Corporation, either expressly, or by estoppel. SeqScape Software has not undergone specific developmental validation for human identification applications. Human identification laboratories that choose to use SeqScape Software. For data analysis should perform their own developmental validation studies. Applera, Applied Biosystems, AB (Design), BigDye, GeneMapper, MicroSeq, and SeqScape are registered trademarks and, KB, Variant Reporter, VariantSEQr and XTerminator are trademarks of the Applera Corporation or its subsidiaries in the US and/or certain other countries. Windows is a registered trademark of Microsoft Corporation. All other trademarks are the sole property of their respective owners. ©2007. Applied Biosystems. All rights reserved. Printed in the USA, 09/2007, Publication #133PB04-01 Headquarters 850 Lincoln Centre Drive | Foster City, CA 94404 USA Phone 650.638.5800 | Toll Free 800.345.5224 www.appliedbiosystems.com
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