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Quantified High-Throughput Screening of Escherichia coli Producing Poly(3-hydroxybutyrate) Based on FACS |
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| Authors: | Jae Hyung Lee Seung Hwan Lee Sung Sun Yim Kyoung-Hee Kang Sang Yup Lee Si Jae Park Ki Jun Jeong |
| Institution: | 1. Department of Chemical and Biomolecular Engineering (BK21 Program), KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea 2. Industrial Biochemicals Research Group, Research Center for Biobased Chemistry, Division of Convergence Chemistry, Korea Research Institute of Chemical Technology, P.O. Box 107, 141 Gajeong-ro, Yuseong-gu, Daejeon, 305-600, Republic of Korea 4. Department of Environmental Engineering and Energy and Department of Energy and Biotechnology, Myongji University, San 38-2, Nam-dong, Cheoin-gu, Yongin-si, Gyeonggido, 449-728, Republic of Korea 3. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Program), Center for Systems and Synthetic Biotechnology, and Institute for the BioCentury, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 305-701, Republic of Korea |
| Abstract: | Here, we report on a highly sensitive method for the detection of P(3HB) accumulation in Escherichia coli cells based on the automated flow cytometry system using fluorescent dyes. E. coli containing P(3HB) were stained with either BODIPY or Nile red fluorescent dye, and their staining properties were analyzed under a variety of conditions. Compared with Nile red, BODIPY was much more sensitive in staining P(3HB) and overall demonstrated a more rapid staining of cells, a greater resistance to photobleaching, and greater cell viability. In addition, we also successfully monitored heterogeneity in P(3HB) accumulation within a cell population using BODIPY staining and flow cytometry. We believe this optimized staining method using BODIPY in combination with screening by high-speed flow cytometer will be helpful in the engineering of host cells toward an enhanced production of bioplastics. |
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