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Localized transfection on arrays of magnetic beads coated with PCR products

Abstract

High-throughput gene analysis would benefit from new approaches for delivering DNA or RNA into cells. Here we describe a simple system that allows any molecular biology laboratory to carry out multiple, parallel cell transfections on microscope coverslip arrays. By using magnetically defined positions and PCR product–coated paramagnetic beads, we achieved transfection in a variety of cell lines. Beads may be added to the cells at any time, allowing both spatial and temporal control of transfection. Because the beads may be coated with more than one gene construct, the method can be used to achieve cotransfection within single cells. Furthermore, PCR-generated mutants may be conveniently screened, bypassing cloning and plasmid purification steps. We illustrated the applicability of the method by screening combinatorial peptide libraries, fused to GFP, to identify previously unknown cellular localization motifs. In this way, we identified several localizing peptides, including structured localization signals based around the scaffold of a single C2H2 zinc finger.

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Figure 1: Magnetically defined transfection arrays.
Figure 2: Confocal microscopy images of selected clones from the colony-PCR bead-transfection screen of the EGFP peptide libraries.

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Acknowledgements

M.I. was supported by an International Research Fellowship from the Wellcome Trust, UK. M.I.S. was funded by a Fundacion Carolina Postdoctoral Fellowship. We would like to thank G. DeCarcer Diez and K. Michalodimitrakis for help in the design of the transfection chamber and D. Megias Vazquez for assistance with confocal imaging.

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Correspondence to Mark Isalan.

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Isalan, M., Santori, M., Gonzalez, C. et al. Localized transfection on arrays of magnetic beads coated with PCR products. Nat Methods 2, 113–118 (2005). https://doi.org/10.1038/nmeth732

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