Introduction
LongTrace:Box™ is a new DNA sequencing system for improving the read length of sequencing traces collected on the ABI 3730, 3730xl, 3100 and 3130 DNA sequencers. The LongTrace:Box system offers the following features:
- Up to 30% more KB Q20+ bases per trace, with an overall average improvement of 10% to 20%.
- Compatible with trace collected on the ABI 3730, 3130, and 3100 series DNA sequencers.
- An all-in-one system with an easy to use graphical user interface that makes it simple to integrate into existing sequencing workflows and LIMS systems.
LongTrace:Box DNA sequencing system
Current state of the art capillary DNA sequencers, such as the ABI 3730 or 3130 series, generate reads that can easily exceed 1000 bases. Unfortunately, the reads often display low quality and
high base call error rates in the later part of the trace (ie post 850 bases).
LongTrace:Box uses a novel approach to extract more high quality base calls from the poorly resolved end regions. The LongTrace software reprocesses the raw data contained within each ABI trace file
using a series of highly sophisticated algorithms developed by Nucleics. This enables the ABI KB basecaller to make significantly better base and quality calls. By combining this raw data reprocessing
with the KB basecaller in a single hardware system, the LongTrace:Box is able to provide a completely seamless method to increase sequencing read lengths without not requiring any changes to current
production protocols. An example of LongTrace processing is shown in Figure 1.
Figure 1. The KB v1.2 derived base and quality calls from an example ABI 3730xl 50cm capillary array sequence. A. The KB base called trace before LongTrace processing. B. The same KB base called trace after LongTrace processing. Note that the trace region shown is at base 1000.
The LongTrace:Box system provides a number of advantages. First, as the LongTrace:Box is a complete system all trace reprocessing can be performed within the facility thus alleviating any possible privacy concerns user may have with our ftp server based LongTrace service. Second, as the output of LongTrace:Box is a standard KB trace file, the traces are from the end users perspective, indistinguishable from a standard KB file.
Features & Benefits of LongTrace:Box
Simple to use
The LongTrace:Box DNA sequencing system is simple to both set up and use. The system is based around a custom Apple Intel Mac Mini running Windows XP Professional. This allows the system to be accessed
using the standard Windows file share network.
The LongTrace:Box also comes with a graphical administration software (Auto LongTrace™) that allows facility manager to automate the LongTrace processing of ABI files located anywhere on the network.
In addition, the Auto LongTrace software provides a series of tab delimited processing reports for both the final end users and facility managers. The online Auto
LongTrace manual provides an overview of the features of the Auto LongTrace software.
Increases trace read length and reduces errors
LongTrace provides the user with significantly longer and higher quality reads (Figures 1 & 2). These improved read lengths mean that fewer sequencing reactions need be performed to complete a sequencing project, thus saving the investigator both time and money.
Figure 2. Distribution of read length improvement after LongTrace processing. Results from 3449 traces generated on an ABI3730xl 36cm capillary array.
Increases DNA sequencing throughput
LongTrace:Box reprocessing of ABI 36cm capillary arrays traces provides read lengths equivalent to that obtained from standard 50cm arrays traces. Since standard 36cm array runs take only half the time of standard 50cm array runs, the effective sequencer throughput can be doubled. This enables facilities to avoid purchasing additional sequencing instruments as demand increases. In addition, Nucleics has developed a modified 36cm array run module that allows up to 28 runs per day (~51 min total run time). This optimized run module provides the same read length potential as the standard 2 hour 50cm array module, effectively permitting the total sequencer throughput to be increased by up to 130%.
The improved read lengths provided by LongTrace:Box increase sequence throughput and significant lower the overall per base cost. In addition, these longer read lengths enable sequencing projects (e.g. whole-genome, cDNA clones, etc) to be completed at lower sequence coverage levels, thus offering further cost and time savings.
The LongTrace:Box system is available at three throughput levels (5000, 10,000 or 25,000 processing per month) with pricing ranging from 6.4 to 16 cents per trace.
For further information on LongTrace:Box please or download the LongTrace:Box overview pdf or view the Auto LongTrace users manual.