Microdroplet enabled cultivation of single yeast cells correlates with bulk growth and reveals subpopulation phenomena

Liu, Hangrui; Xu, Xin; Peng, Kang; Zhang, Yuxin; Jiang, Lianmei; Williams, Thomas; Paulsen, Ian T.; Piper, James A.; Li, Ming

doi:10.1002/bit.27591

Cited by 20 publications

(14 citation statements)

References 60 publications

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“…Alternatively, DE can be used as an intermediate template to produce microgels in aqueous solution, avoiding the use of oil and the extra washing step. ,,, Using the microfluidic device described here, we have created monodisperse gelatin-in-oil-in-water DEs with different sizes and compositions. Gelatin is a cross-linked natural protein with thermally responsive sol–gel transition properties and is commonly used in food and pharmaceutical industries …”

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…The procedures for the preparation of GFP-tagged Saccharomyces cerevisiae (S. cerevisiae) strain (CEN.PK2-1C-GFP), determination of concentration, and in vitro culture were detailed in our previous study. 47 GFP-tagged S. cerevisiae cells at four different concentrations: 5 × 10 6 , 1 × 10 7 , 2 × 10 7 , and 8 × 10 7 cells/mL were used in the inner phase. A cell concentration of 1 × 10 7 cells/mL was used for the screening of single-cell growth in DEs.…”

Section: ■ Methods and Materialsmentioning

confidence: 99%

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Rapid, Simple, and Inexpensive Spatial Patterning of Wettability in Microfluidic Devices for Double Emulsion Generation

Liu

Piper

2021

Anal. Chem.

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Water-in-oil-in-water (w/o/w) double emulsion (DE) encapsulation has been widely used as a promising platform technology for various applications in the fields of food, cosmetics, pharmacy, chemical engineering, materials science, and synthetic biology. Unfortunately, DEs formed by conventional emulsion generation approaches in most cases are highly polydisperse, making them less desirable for quantitative assays, controlled biomaterial synthesis, and entrapped ingredient release. Microfluidic devices can generate monodisperse DEs with controllable size, morphology, and production rate, but these generally require multistep fabrication processes and use of different solvents or bulky external instrumentation to pattern channel wettability. To overcome these limitations, we propose a rapid, simple, and inexpensive method to spatially pattern wettability in microfluidic devices for the continuous generation of monodisperse DEs. This is achieved by applying corona−plasma treatment to a select zone of the microchannel surface aided by a custom-designed corona resistance microchannel to strictly confine the plasma-treatment zone in a single polydimethylsiloxane (PDMS) microfluidic device. The properties of PDMS channel surfaces and key microchannel regions for DE generation are characterized under different levels of treatment. The size, shell thickness, and number of inner cores of generated DEs are shown to be highly controllable by tuning the phase flow rate ratios. Using DEs as templates, we successfully achieve a one-step generation and collection of gelatin microgels. Additionally, we demonstrate the biological capability of generated DEs by flow cytometric screening of the encapsulation and growth of yeast cells within DEs. We expect that the proposed approach will be widely used to create microfluidic devices with more complex wettability patterns.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: ■ Methods and Materialsmentioning

confidence: 99%

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Rapid, Simple, and Inexpensive Spatial Patterning of Wettability in Microfluidic Devices for Double Emulsion Generation

Liu

Piper

2021

Anal. Chem.

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“…The study of a uniform S. Cerevisiae population enables us to better understand the underling molecular mechanisms of producing valuable compounds (e.g., enzymes, pharmaceutical proteins, and organic acids), as well as biological processes in the human body, diseases, and cancers (e.g., brain and nervous system development and Parkinson’s disease , ). Moreover, synchronization of cell cycle is critical for many biological experiments as the heterogeneity in cell cycle often leads to misinterpretation of the experimental data . Separation and synchronization of S. Cerevisiae by shape can enrich cell populations for a single stage, improving the examination of the connection of genes and proteins with the functions that they provide to cells.…”

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confidence: 99%

Separation and Enrichment of Yeast Saccharomyces cerevisiae by Shape Using Viscoelastic Microfluidics

Liu

Yuan

et al. 2020

Anal. Chem.

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Yeast Saccharomyces cerevisiae (S. Cerevisiae) is one of the most attractive microbial species used for industrial production of value-added products and is an important model organism to understand the biology of the eukaryotic cells and humans. S. Cerevisiae has different shapes, such as spherical singlets, budded doublets, and clusters, corresponding to phases of the cell cycle, genetic, and environmental factors. The ability to obtain high-purity populations of uniform-shaped S. Cerevisiae cells is of significant importance for a wide range of applications in basic biological research and industrial processes. In this work, we demonstrate shape-based separation and enrichment of S. Cerevisiae using a coflow of viscoelastic and Newtonian fluids in a straight rectangular microchannel. Due to the combined effects of lift inertial and elastic forces, this label-free and continuous separation arises from shape-dependent migration of cells from the Newtonian to the non-Newtonian viscoelastic fluid. The lateral position of S. Cerevisiae cells with varying morphologies is found to be dependent on cell major axis. We also investigate the effects of sheath and sample flow rate, poly(ethylene oxide) (PEO) concentration and channel length on the performance of the viscoelastic microfluidic device for S. Cerevisiae enrichment and separation by shape. Moreover, the separation efficiency, cell extraction yield, and cell viability after sorting operations are studied.

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“…GMDs are small spheres of hydrogel in which few or even single cells can be encapsulated [12]. The small volume of GMDs makes them highly permeable, which allows for communication among cells and the diffusion of cellular metabolites [12,13]. Culturing marine sponge cells presents some unique challenges, for example, marine sponges require high salinities which can prevent the hydrogels from solidifying properly.…”

Section: Introductionmentioning

confidence: 99%

3-D Culture of Marine Sponge Cells for Production of Bioactive Compounds

et al. 2021

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Production of sponge-derived bioactive compounds in vitro has been proposed as an alternative to wild harvest, aquaculture, and chemical synthesis to meet the demands of clinical drug development and manufacture. Until recently, this was not possible because there were no marine invertebrate cell lines. Recent breakthroughs in the development of sponge cell lines and rapid cell division in improved nutrient media now make this approach a viable option. We hypothesized that three-dimensional (3-D) cell cultures would better represent how sponges function in nature, including the production of bioactive compounds. We successfully cultured sponge cells in 3-D matrices using FibraCel® disks, thin hydrogel layers, and gel microdroplets (GMDs). For in vitro production of bioactive compounds, the use of GMDs is recommended. Nutrients and sponge products rapidly diffuse into and out of the 3-D matrix, the GMDs may be scaled up in spinner flasks, and cells and/or secreted products can be easily recovered. Research on scale-up and production is in progress in our laboratory.

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Microdroplet enabled cultivation of single yeast cells correlates with bulk growth and reveals subpopulation phenomena

Cited by 20 publications

References 60 publications

Rapid, Simple, and Inexpensive Spatial Patterning of Wettability in Microfluidic Devices for Double Emulsion Generation

Rapid, Simple, and Inexpensive Spatial Patterning of Wettability in Microfluidic Devices for Double Emulsion Generation

Separation and Enrichment of Yeast Saccharomyces cerevisiae by Shape Using Viscoelastic Microfluidics

3-D Culture of Marine Sponge Cells for Production of Bioactive Compounds

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