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Issue 106, 2014
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Capture and culturing of single microalgae cells, and retrieval of colonies using a perforated hemispherical microwell structure

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Abstract

A perforated hemispherical microwell structure is shown to efficiently capture single Chlamydomonas reinhardtii (C. reinhardtii) cells, culture them to form colonies, and retrieve these colonies to serve as seeds for large-scale cultivation. This solution-phase formation and recovery of colonies could overcome the tedious and time-consuming process of selecting colonies from a solid-phase agar plate. The fabricated microdevice was composed of three layers: a top layer consisting of a cell solution for injection and recovery of a microalgal solution, a hemispherical perforated microwell array in the middle, and a bottom layer in which the solution is manipulated by controlling the hydrodynamic force. The microalgal (wild type and hygromycin B-resistant mutant) cells loaded in the top layer rapidly diffused into the microwell holes, and individual such cells were captured with high efficiency (>90%) and within 1 min by applying a withdraw mode in the bottom layer. Single-cell-based cultivation in a medium containing hygromycin B was then performed to generate colonies in the hemispherical microwell. While the wild type cells died, mutant cells resistant to hygromycin B survived well and grew into a colony within 2 days. The produced colonies in the microwells were recovered by applying a release mode in the bottom layer, so that a hydrodynamic force was exerted vertically to push out the colonies through the outlet in 10 s. The recovered cells were cultured on a large scale in medium by using a flask. The recovered C. reinhardtii was confirmed as a hygromycin B-resistant mutant by identifying the hygromycin gene in the polymerase chain reaction (PCR). The microdevice described here could in solution perform single-cell capture, colony formation, and retrieval of colonies for further large-scale cultivation, which could replace tedious and time-consuming solid-phase agar plate processes with a 7-fold reduction in the duration of the process.

Graphical abstract: Capture and culturing of single microalgae cells, and retrieval of colonies using a perforated hemispherical microwell structure

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Publication details

The article was received on 03 Sep 2014, accepted on 31 Oct 2014 and first published on 31 Oct 2014


Article type: Paper
DOI: 10.1039/C4RA09730K
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RSC Adv., 2014,4, 61298-61304

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    Capture and culturing of single microalgae cells, and retrieval of colonies using a perforated hemispherical microwell structure

    J. S. Choi, S. Bae, K. H. Kim, J. Y. H. Kim, S. J. Sim and T. S. Seo, RSC Adv., 2014, 4, 61298
    DOI: 10.1039/C4RA09730K

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