Melt‐Encoded‐Tags for Expanded Optical Readout in Digital PCR (METEOR‐dPCR) Enables Highly Multiplexed Quantitative Gene Panel Profiling

Author:

Liu Dong Dong1ORCID,Muliaditan Daniel23,Viswanathan Ramya12,Cui Xu2,Cheow Lih Feng12ORCID

Affiliation:

1. Institute for Health Innovation and Technology National University of Singapore Singapore 117599 Singapore

2. Department of Biomedical Engineering Faculty of Engineering National University of Singapore Singapore 117583 Singapore

3. Genome institute of Singapore Agency for Science Technology and Research Singapore 138672 Singapore

Abstract

AbstractDigital PCR (dPCR) is an important tool for precise nucleic acid quantification in clinical setting, but the limited multiplexing capability restricts its applications for quantitative gene panel profiling. Here, this work describes melt‐encoded‐tags for expanded optical readout in digital PCR (METEOR‐dPCR), a simple two‐step assay that enables simultaneous quantification of a large panel of arbitrary genes in a dPCR platform. Target genes are quantitatively converted into DNA tags with unique melting temperatures through a ligation approach. These tags are then counted and distinguished by their melt‐curve profiles on a dPCR platform. A multiplexing capacity of M^N, where M is the number of resolvable melting temperature and N is the number of fluorescence channel, can be achieved. This work validates METEOR‐dPCR with simultaneous DNA copy number profiling of 60 targets using dPCR in cancer cells, and demonstrates its sensitivity for estimating tumor fraction in mixed tumor and normal DNA samples. The rapid, quantitative, and highly multiplexed METEOR‐dPCR assay will have wide appeal for many clinical applications.

Funder

Ministry of Education

Publisher

Wiley

Subject

General Physics and Astronomy,General Engineering,Biochemistry, Genetics and Molecular Biology (miscellaneous),General Materials Science,General Chemical Engineering,Medicine (miscellaneous)

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