BEAMing: single-molecule PCR on microparticles in water-in-oil emulsions

@article{Diehl2006BEAMingSP,
  title={BEAMing: single-molecule PCR on microparticles in water-in-oil emulsions},
  author={Frank Diehl and Meng Li and Yiping He and Kenneth W. Kinzler and Bert Vogelstein and Devin Dressman},
  journal={Nature Methods},
  year={2006},
  volume={3},
  pages={551-559}
}
The most important biotechnological advances made in the 20th century involved methods that converted single DNA molecules into populations of identical DNA molecules. The first wave of techniques used cells (cloning)1 and the second used PCR2. Cloning was advantageous because the populations arising from individual molecules were inherently separated. In contrast, each template required individual compartments (tubes) for PCR-based methods if separate products were desired. Emulsion PCR… 
Solid-phase PCR in a picowell array for immobilizing and arraying 100,000 PCR products to a microscope slide.
TLDR
Compared to widely established emulsion based PCR (emPCR) approaches, leading to PCR products immobilized onto bead surfaces in a highly parallel manner, the novel technique results in direct spatial registration of immobilized PCR products in a microarray format.
Optimization of On-Bead Emulsion Polymerase Chain Reaction Based on Single Particle Analysis
TLDR
It was found that increasing the polymerase concentration 20-fold in ePCR over recommended concentrations for conventional PCR was necessary to obtain sufficient PCR products and Surprisingly, dramatical increases in the concentrations of reverse primer and nucleotides recommended in literature gave no measurable change in outcome.
An Emulsion System Based on a Chip Polymerase Chain Reaction
TLDR
A novel method for detecting many DNA fragments through efficient amplification by using an emulsion system based on “on-chip” PCR instead of conventional multiplex polymerase chain reaction (PCR).
Chemoselective Coupling Preserves the Substrate Integrity of Surface-Immobilized Oligonucleotides for Emulsion PCR-Based Gene Library Construction.
TLDR
This work describes covalent bead functionalization with primers via either azide-alkyne click chemistry or Michael addition, a simpler, chemical approach to primer immobilization that may spur more economical library preparation for high-throughput sequencing and enable more complex surface elaboration for in vitro evolution.
Real-time PCR of single bacterial cells on an array of adhering droplets.
TLDR
An inexpensive chip-level device that is compatible with a commercial real-time PCR thermal cycler to perform quantitative PCR for single bacterial cells and showed that the device is capable of genetic analysis at single bacterial cell level with C(q) standard deviation less than 1.05 cycles.
Single-molecule emulsion PCR in microfluidic droplets
TLDR
The important technical breakthroughs of microfluidic droplet PCR in the past five years and their applications to single-molecule amplification and analysis, such as high-throughput screening, next generation DNA sequencing, and quantitative detection of rare mutations are reviewed.
An emulsion digital PCR quantitative method based on microbeads and micropillar array chip
  • Zule Cheng, Kun Wang, Hongju Mao
  • Engineering
    2017 19th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS)
  • 2017
BEAMing (beads, emulsion, amplification, and magnetics) is a high sensitive emulsion digital PCR (polymerase chain reaction) technology. Target beads obtained via BEAMing can be used to determine the
Agarose droplet microfluidics for highly parallel and efficient single molecule emulsion PCR.
TLDR
An agarose droplet method was developed for highly parallel and efficient single molecule emulsion PCR that allows high throughput generation of uniform droplets and enables high PCR efficiency, making it a promising platform for many single copy genetic studies.
...
...

References

SHOWING 1-10 OF 17 REFERENCES
PCR amplification from single DNA molecules on magnetic beads in emulsion: application for high-throughput screening of transcription factor targets
TLDR
A novel method of genetic library construction on magnetic microbeads based on solid-phase single-molecule PCR in a fine and robust water-phase compartment formed in water-in-oil emulsions, which is a powerful tool for analyzing the transcription network on a genomewide scale.
Transforming single DNA molecules into fluorescent magnetic particles for detection and enumeration of genetic variations
TLDR
This approach is called BEAMing on the basis of four of its principal components (beads, emulsion, amplification, and magnetics) and can be used for the identification of rare mutations as well as to study variations in gene sequences or transcripts in specific populations or tissues.
Solid phase DNA amplification: a Brownian dynamics study of crowding effects.
TLDR
The results indicate that the steric interaction between molecules leads to a decreased duplication probability for molecules in the center of a colony and to an outward leaning for the molecules on the perimeter, which result in slower amplification (compared to solution PCR) and indicate that steric interactions alone can explain the loss of the exponential growth of the number of molecules in an SPA experiment.
Development of a microchamber array for picoliter PCR.
TLDR
The lower volume limit for PCR was investigated using various sizes of microchambers, and the system was improved in order to take up the PCR product.
Genome sequencing in microfabricated high-density picolitre reactors
TLDR
A scalable, highly parallel sequencing system with raw throughput significantly greater than that of state-of-the-art capillary electrophoresis instruments with 96% coverage at 99.96% accuracy in one run of the machine is described.
Man-made cell-like compartments for molecular evolution
TLDR
It is demonstrated the linkage of genotype to phenotype in man-made compartments using a model system and a selection for target-specific DNA methylation was based on the resistance of the product (methylated DNA) to restriction digestion.
Accurate Multiplex Polony Sequencing of an Evolved Bacterial Genome
We describe a DNA sequencing technology in which a commonly available, inexpensive epifluorescence microscope is converted to rapid nonelectrophoretic DNA sequencing automation. We apply this
Monodisperse Double Emulsions Generated from a Microcapillary Device
TLDR
It is shown that the droplet size can be quantitatively predicted from the flow profiles of the fluids, which makes this a flexible and promising technique.
...
...