Long Range PCR Amplification and Amplicon Normalization

The SequalPrep™ (Sequence Equalization Sample Prep) System is the first complete long PCR enrichment and amplicon normalization system that provides a highly efficient enrichment solution to meet NGS re-sequencing throughput capabilities. The system offers the dual benefit of more enhanced and consistent coverage of your region(s) of interest with “factory-proof” long-range PCR amplification and a simple, one-step amplicon equalization solution in a 96 well format. The system is comprised of the SequalPrep™ Long PCR Kit with dNTPs and the SequalPrep™ Normalization Plate.

SequalPrep™ Long Range PCR Kit with dNTPs

Figure 1 Success rate of six long PCR kit vendors. Invitrogen's SequalPrep™ Long PCR Kit with dNTPs was the only kit that successfully amplified all six 10 kB amplicons without the need for additional optimization.

Long range PCR amplification success with minimal optimization.

The SequalPrepTM Long PCR Kit with dNTPs provides the highest amplification success rate of any commercially available kit tested across multiple organisms and platforms for amplicons up to 20 kB with little to no optimization required (Figure 1).

PCR amplification of the target region of interest is the traditional method used to enrich areas of the genome upstream of medical re-sequencing to improve PCR sequencing efficiency for studies of small gene families. To take advantage of NGS processing capabilities higher throughput enrichment methods are required. A simple solution to reduce PCR complexity is to increase the size of each amplicon. Unfortunately, efficiency gains from long range PCR have been offset by lower amplification success rates offered by available commercial solutions.

After intensive optimization of numerous PCR enhancement technologies and extensive field testing of human and mouse targets, the SequalPrep™ Long PCR Kit with dNTPs provides the highest amplification success rate for long range PCR targets.

SequalPrep™ Long PCR Kit with dNTPs Frequently-Asked-Questions

SequalPrep™ Normalization Plate (96)

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Figure 2 Comparison of conventional normalization and SequalPrep™ normalization methods. Invitrogen's SequalPrep™ Normalization Plate provides a convenient, one-step amplicon purification and normalization solution in a 96 well format.

Amplicon purification and normalization in just one simple step

Based on a simple, yet elegant binding capacity saturation method, the SequalPrep™ Normalization Plate offers an easily automated solution for amplicon equalization and downstream consistency of sequencing coverage (Figure 2).

Due to the variable efficiencies in amplification reactions of target regions, amplicon yield can vary significantly post PCR amplification. Variation in amplicon yield can lead to lower depth of coverage for amplicons in lower abundance. Conventional methods used to ensure equal and consistent representation are laborious due to the numerous sample manipulation steps involved in purification and quantification.

The SequalPrep™ Normalization Plate provides a convenient, one-step amplicon purification and normalization solution in a 96 well format.

SequalPrep™ Long PCR Kit with dNTPs Frequently-Asked-Questions

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