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Analytical and Bioanalytical Chemistry

, Volume 411, Issue 10, pp 2161–2168 | Cite as

Combination of a flow cytometric bead system with 16S rRNA-targeted oligonucleotide probes for bacteria detection

  • Yan Zeng
  • Dun ZhangEmail author
  • Peng QiEmail author
Research Paper
  • 72 Downloads

Abstract

Here we report a bacteria detection method based on a flow cytometric bead system and 16S rRNA-targeted oligonucleotide probes. Polymerase chain reaction (PCR) was first used to acquire bacterial DNA including bacteria-specific sequences. Half of the resulting target DNA was then captured by a capture probe immobilized on a magnetic microbead (MB) surface. The other half of the target DNA was hybridized with a fluorescence-labeled signal probe. In this manner, a sandwich DNA hybridization involving a MB-based capture probe, the target DNA, and a signal probe was realized. The MB carriers modified with reporter dye were analyzed one by one by flow cytometry through a capillary. Using PCR amplicons and this flow cytometric bead system, a detection limit of 180 cfu mL−1 was achieved, along with high selectivity that permitted the discrimination of different targets when challenged with control bacteria targets and multiplexing capabilities that enabled the simultaneous detection of two kinds of bacteria. Given these advantages, the developed method can be used for the highly sensitive and specific PCR amplicon analysis of DNA extracted from a fresh bacterial culture, as well as multiplex target analysis.

Graphical abstract

The flow cytometric bead system with 16S rRNA-targeted oligonucleotide probes for bacteria detection developed in this work. This system is highly specific and sensitive, with a detection limit of 180 cfu mL−1 bacteria.

Keywords

Bacteria detection Flow cytometry Magnetic beads 16S rRNA 

Notes

Acknowledgements

We gratefully acknowledge the support provided by the National Natural Science Foundation of China (41876101) and project funding from the China Postdoctoral Science Foundation (2016 M602199), the Young Elite Scientists Sponsorship Program by CAST (2018QNRC001), the Applied Basic Research Program of Nantong (GY12017005), and the AoShan Talent Program supported by the Qingdao National Laboratory for Marine Science and Technology.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

216_2019_1651_MOESM1_ESM.pdf (414 kb)
ESM 1 (PDF 414 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Key Laboratory of Marine Environmental Corrosion and Biofouling, Institute of OceanologyChinese Academy of SciencesQingdaoChina
  2. 2.Center for Ocean MegascienceChinese Academy of SciencesQingdaoChina
  3. 3.Open Studio for Marine Corrosion and ProtectionQingdao National Laboratory for Marine Science and TechnologyQingdaoChina

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