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A robust protocol to increase NimbleGen SeqCap EZ multiplexing capacity to 96 samples.

Ilse M van der Werf1, R Frank Kooy1, Geert Vandeweyer1

  • 1Department of Medical Genetics, University of Antwerp, Antwerp, Belgium.

Plos One
|April 16, 2015
PubMed
Summary
This summary is machine-generated.

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Researchers enhanced the NimbleGen SeqCap EZ protocol to multiplex up to 96 samples using dual-indexing. This cost-effective method improves targeted gene panel sequencing by increasing sample capacity and optimizing pooling factors.

Area of Science:

  • Genomics
  • Molecular Biology
  • Bioinformatics

Background:

  • Targeted gene panel sequencing is crucial for genetic studies and disease research.
  • The NimbleGen SeqCap EZ Choice kit facilitates targeted enrichment but is limited by single-indexing, restricting multiplexing to 24 samples.
  • High data output per sample occurs with small target sizes and limited multiplexing.

Purpose of the Study:

  • To enhance the NimbleGen SeqCap EZ protocol for increased sample multiplexing.
  • To optimize cost-efficiency for custom target panel sequencing.
  • To provide tools for evaluating capture efficiency in custom panels.

Main Methods:

  • Incorporation of Illumina dual-indexing adapters into the NimbleGen SeqCap EZ protocol.
  • Investigation of higher pre-enrichment pooling factors (up to 12 samples).

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  • Development of a reporting tool for capture efficiency evaluation.

    • Main Results:

    • Successfully multiplexed up to 96 samples using dual-indexing.
    • Demonstrated that pre-enrichment pooling up to 12 samples does not compromise data quality.
    • Developed a functional reporting tool for custom panel capture efficiency.

    Conclusions:

    • The modified NimbleGen SeqCap EZ protocol significantly increases sample multiplexing capacity.
    • The enhanced protocol offers a more cost-effective solution for targeted gene panel sequencing.
    • The developed tools aid in the efficient design and evaluation of custom sequencing panels.