Liquid marbles as biochemical reactors for the polymerase chain reaction

Sreejith, Kamalalayam Rajan; Gorgannezhad, Lena; Jin, Jing; Ooi, Chin Hong; Stratton, Helen; Dao, Dzung Viet; Nguyen, Nam‐Trung

doi:10.1039/c9lc00676a

Cited by 52 publications

(36 citation statements)

References 54 publications

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“…Next, the fluorescent intensity values of the samples inside the microchambers of the device were normalised. Normalisation of the fluorescent intensities was achieved by the following formula [27]:…”

Section: Simultaneous Detection Of Microbial Faecal Source Tracking (Mst) Markers Using Microfluidic Devicementioning

confidence: 99%

Microfluidic Array Chip for Parallel Detection of Waterborne Bacteria

et al. 2019

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The polymerase chain reaction (PCR) is a robust technique used to make multiple copies of a segment of DNA. However, the available PCR platforms require elaborate and time-consuming operations or costly instruments, hindering their application. Herein, we introduce a sandwiched glass–polydimethylsiloxane (PDMS)–glass microchip containing an array of reactors for the real-time PCR-based detection of multiple waterborne bacteria. The PCR solution was loaded into the array of reactors in a single step utilising capillary filling, eliminating the need for pumps, valves, and liquid handling instruments. Issues of generating and trapping bubbles during the loading chip step were addressed by creating smooth internal reactor surfaces. Triton X-100 was used to enhance PCR compatibility in the chip by minimising the nonspecific adsorption of enzymes. A custom-made real-time PCR instrument was also fabricated to provide thermal cycling to the array chip. The microfluidic device was successfully demonstrated for microbial faecal source tracking (MST) in water.

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Section: Simultaneous Detection Of Microbial Faecal Source Tracking (Mst) Markers Using Microfluidic Devicementioning

confidence: 99%

Microfluidic Array Chip for Parallel Detection of Waterborne Bacteria

et al. 2019

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“…The major issue of using a conventional liquid marble for polymerase chain reaction is the evaporation of the PCR mixture at elevated temperatures. We have previously developed a simple method to reduce evaporation of the PCR mixture in liquid marbles and successfully conducted PCR in a liquid marble coated with polytetrafluoroethylene (PTFE) powder [32]. However, the sample volume used in the experiment was relatively large (50 µl) and the PCR mixture could only last for 9 to 10 thermal cycles due to evaporation of PTFE-coated liquid marbles [33].…”

Section: Introductionmentioning

confidence: 99%

Core-Shell Beads Made by Composite Liquid Marble Technology as A Versatile Microreactor for Polymerase Chain Reaction

et al. 2020

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Over the last three decades, the protocols and procedures of the DNA amplification technique, polymerase chain reaction (PCR), have been optimized and well developed. However, there have been no significant innovations in processes for sample dispersion for PCR that have reduced the amount of single-use or unrecyclable plastic waste produced. To address the issue of plastic waste, this paper reports the synthesis and successful use of a core-shell bead microreactor using photopolymerization of a composite liquid marble as a dispersion process. This platform uses the core-shell bead as a simple and effective sample dispersion medium that significantly reduces plastic waste generated compared to conventional PCR processes. Other improvements over conventional PCR processes of the novel dispersion platform include increasing the throughput capability, enhancing the performance and portability of the thermal cycler, and allowing for the contamination-free storage of samples after thermal cycling.

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“…Due to the small dimension of a liquid marble, the reagent consumption can be reduced [40,41], particularly as for chemical and biological applications [42]. More recently, Sreejith et al [43] used liquid marbles as reactors for DNA amplification. Polytetrafluoroethylene (PTFE) was used as the hydrophobic coating.…”

Section: Introductionmentioning

confidence: 99%

Core-Shell Beads as Microreactors for Phylogrouping of E. coli Strains

et al. 2020

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Multiplex polymerase chain reaction (PCR) is an effective tool for simultaneous detection of target genes. Nevertheless, their use has been restricted due to the intrinsic interference between primer pairs. Performing several single PCRs in an array format instead of a multiplex PCR is a simple way to overcome this obstacle. However, there are still major technical challenges in designing a new generation of single PCR microreactors with a small sample volume, rapid thermal cycling, and no evaporation during amplification. We report a simple and robust core-shell bead array for a series of single amplifications. Four core-shell beads with a polymer coating and PCR mixture were synthesized using liquid marble formation and subsequent photo polymerization. Each bead can detect one target gene. We constructed a customised system for thermal cycling of these core-shell beads. Phylogrouping of the E. coli strains was carried out based on the fluorescent signal of the core-shell beads. This platform can be a promising alternative for multiplex nucleic acid analyses due to its simplicity and high throughput. The platform reported here also reduces the cycling time and avoids evaporation as well as contamination of the sample during the amplification process.

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Liquid marbles as biochemical reactors for the polymerase chain reaction

Abstract: Liquid marbles can serve as a biochemical reactor for the polymerase chain reaction, eliminating the conventional single use plastic reaction vial.

Cited by 52 publications

References 54 publications

Microfluidic Array Chip for Parallel Detection of Waterborne Bacteria

Microfluidic Array Chip for Parallel Detection of Waterborne Bacteria

Core-Shell Beads Made by Composite Liquid Marble Technology as A Versatile Microreactor for Polymerase Chain Reaction

Core-Shell Beads as Microreactors for Phylogrouping of E. coli Strains

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