High-Throughput Droplet Digital PCR System for Absolute Quantitation of DNA Copy Number
Высокопроизводительная система капельной цифровой ПЦР для абсолютного количественного определения числа копий ДНК
2011-10-28
SCID: 54.1/kda25f4h
Discuss with AI
absolute DNA quantitationcell-free fetal DNAcopy number variationdroplet digital PCRrare allele detection
Figures from the paper
Abstract (AI)
Digital PCR enables the absolute quantitation of nucleic acids in a sample. The lack of scalable and practical technologies for digital PCR implementation has hampered the widespread adoption of this inherently powerful technique. Here we describe a high-throughput droplet digital PCR (ddPCR) system that enables processing of ~2 million PCR reactions using conventional TaqMan assays with a 96-well plate workflow. Three applications demonstrate that the massive partitioning afforded by our ddPCR system provides orders of magnitude more precision and sensitivity than real-time PCR. First, we show the accurate measurement of germline copy number variation. Second, for rare alleles, we show sensitive detection of mutant DNA in a 100,000-fold excess of wildtype background. Third, we demonstrate absolute quantitation of circulating fetal and maternal DNA from cell-free plasma. We anticipate this ddPCR system will allow researchers to explore complex genetic landscapes, discover and validate new disease associations, and define a new era of molecular diagnostics.
Key Findings
1
A high-throughput droplet digital PCR system processes approximately 2 million PCR reactions using conventional TaqMan assays in a 96-well plate workflow.
2
Massive sample partitioning provides orders-of-magnitude greater precision and sensitivity than real-time PCR.
3
The platform enables absolute quantitation of circulating fetal and maternal cell-free DNA in plasma.
4
The system accurately measures germline DNA copy-number variation.
5
The system detects rare mutant alleles in a 100,000-fold excess of wild-type DNA.
Research Object
High-throughput droplet digital PCR (ddPCR) system and the DNA samples analyzed with it, including germline, rare mutant, and circulating fetal and maternal DNA
Research Subject
Absolute DNA copy-number quantitation and the system’s precision and sensitivity for measuring copy-number variation, detecting rare mutant alleles, and quantifying cell-free fetal and maternal DNA
Publication Details
Publication Date
2011-10-28
Journal
Publisher
ISSN
Open access PDF
Access Type
Author Information
Download PDF
Subscribe to digest