Ampure® Purification System

Wednesday, February 11, 2004

AMPure is a high recovery, SPRI™-based PCR clean up system capable of purifying amplicons as small as 100bp. AMPure yields high quality PCR amplicons using a simple, three-step protocol requiring no centrifugation or filtration steps. The protocol can be easily automated or performed manually in both 96 and 384-well formats.

EFFICIENT PCR PURIFICATION
AMPure provides:

Efficient removal of unincorporated dNTPs, primers and salts

High recovery of small and large amplicons

High quality PCR amplicons

FLEXIBLE PURIFICATION
AMPure is flexible enough to be performed in a variety of formats for an array of applications:
PLATFORMS: APPLICATIONS:

96-well

PCR Sequencing

384-well

cDNA labeling

Fully automated

Rolling Circle Amplification

Manual

EFFECTIVE CLEAN UP
AMPure removes unincorporated primers, dNTPs and salts used during PCR amplification. These products can interfere with downstream applications. AMPure routinely has recovery rates greater than 90% for amplicons larger than 200bp and greater than 85% for smaller PCR amplicons, such as the 165bp fragment shown in Figure 2.

HIGH PURITY – HIGH PERFORMANCE
AMPure-purified samples are ideal sequencing templates. The table below shows sequencing results from a single 976bp PCR template purified using AMPure or ExoSAP-IT™. The average Phred20 read length and average pass rate for the AMPure-purified samples were 701 and 100% respectively. ExoSAP-IT, an alternative approach of inactivating polymerases and degrading unincorporated primers following PCR thermo-cycling, produces samples with lower purity due to the presence of salts and degraded enzymes and primers. Sequence results following ExoSAP-IT digestions are consistently inferior, with fewer Phred20 bases per read and lower pass rates. Figure 3 exhibits the superior sequencing performance attained using AMPure.

RELIABLE RESULTS
AMPure produces reliable recoveries of both large and small amplicons. Figure 4 shows two gels following AMPure purification of 967 bp and 165 bp PCR fragments.

SUPERIOR PERFORMANCE
AMPure demonstrates excellent recovery rates of both small and large amplicons while maintaining high sample purity. AMPure samples have little to no contamination as shown by the lack of smearing above the bands (Figure 5). In contrast, alternative purification products have decreased recovery rates and/or lower sample purity, which affect the performance of downstream applications such as sequencing.

COMPLETE AMPURE AUTOMATION
The lack of centrifugation and filtration steps allows AMPure protocols to be completely automated while maintaining superior performance. AMPure is fully compatible with most commonly used low, medium and high throughout liquid handling systems. Contact Agencourt for the most up-to-date list of supported platforms.