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| Item Type: | Article |
|---|---|
| Title: | Enrichment of gut microbiome strains for cultivation-free genome sequencing using droplet microfluidics |
| Creators Name: | Pryszlak, A., Wenzel, T., Seitz, K.W., Hildebrand, F., Kartal, E., Cosenza, M.R., Benes, V., Bork, P. and Merten, C.A. |
| Abstract: | We report a droplet microfluidic method to target and sort individual cells directly from complex microbiome samples and to prepare these cells for bulk whole-genome sequencing without cultivation. We characterize this approach by recovering bacteria spiked into human stool samples at a ratio as low as 1:250 and by successfully enriching endogenous Bacteroides vulgatus to the level required for de novo assembly of high-quality genomes. Although microbiome strains are increasingly demanded for biomedical applications, a vast majority of species and strains are uncultivated and without reference genomes. We address this shortcoming by encapsulating complex microbiome samples directly into microfluidic droplets and amplifying a target-specific genomic fragment using a custom molecular TaqMan probe. We separate those positive droplets by droplet sorting, selectively enriching single target strain cells. Finally, we present a protocol to purify the genomic DNA while specifically removing amplicons and cell debris for high-quality genome sequencing. |
| Keywords: | Droplet Microfluidics, Single-Cell Analysis, Genomics, Genome, Gut Microbiome, Functional Analysis, Metagenomics, Bacteria |
| Source: | Cell Reports Methods |
| ISSN: | 2667-2375 |
| Publisher: | Elsevier |
| Volume: | 2 |
| Number: | 1 |
| Page Range: | 100137 |
| Date: | 24 January 2022 |
| Official Publication: | https://doi.org/10.1016/j.crmeth.2021.100137 |
| PubMed: | View item in PubMed |
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