Sanger sequencing, also known as “chain terminator sequencing,” is a method of DNA sequencing that relies on DNA polymerase’s selective incorporation of fluorescent-labeled (bigdye terminator) dideoxy nucleotide chain terminators during in vitro DNA replication for use in a single sequencing reaction. The chain termination PCR produces DNA fragments of varying lengths, each of which ends with a fluorescently labeled dideoxynucleoside. It functions similarly to standard PCR, with one key exception. In contrast to regular PCR, a low ratio of modified nucleotides is introduced along with normal dNTPs. These modified dNTPs are known as dideoxynucleosides (ddNTPs) and have a fluorescent label known as Dye. It quickly became the most used sequencing method for a variety of applications, including de novo sequencing, mutation discovery and confirmation, and resequencing.

After the cycle sequencing reaction, it is necessary to remove contaminants from DNA extension products (e.g., unincorporated fluorescent dyes, residual salts, dNTPs, primers, and enzymes). These impurities, such as dye peaks or “dye blobs,” can frequently interfere with the quality and signal intensity of sequencing data, obscuring sections of the sequencing chromatogram and interfering with the base-calling accuracy of sequencing analysis tools. To clean and purify the sequencing reaction extension items on the market, a number of dye-terminator removal (sequencing clean up or sequencing purification) products are available. However, those sequencing purification protocols are either time-consuming (for example, using a spin column, ethanol precipitation, and SPRI paramagnetic beads) or result in sample loss due to protein precipitation, several transfer stages, and a lengthy process.

BcMag™ One-Step Sequencing Cleanup Kit is specifically designed for fast and efficient purification of the post-Sanger Sequencing reaction. The entire protocol takes only one tube and is complete in less than 5minutes. The magnetic beads are added directly to the finished sequencing reactions and vortexed to capture the impurities (e.g., unincorporated dyes, dNTPs, residual salts, and other interfering components). After vortexing, the beads are magnetically captured, while the clean supernatant can be directly loaded onto a capillary sequencer.

Features and Advantages

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One tube, 3 min protocol, No sample loss

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Reliable results: excellent Long and short fragment recovery, Q20 read length > 800 bases.

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Cost-Effective: Tremendously reduced labor costs and other consumed material such as columns, filters, laborious repeat pipetting, and ethanol.

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High throughput: Compatible with many different automated liquid handling systems.

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Compatible with BigDye Xterminator run modules, e.g., unnecessary to remove the magnetic beads from the tube, the supernatant can be directly loaded onto the capillary sequencer.

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Efficient removal of any dye terminator

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Instruction Manual

MSDS