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Pooled CRISPRi screening of the cyanobacterium Synechocystis sp PCC 6803 for enhanced industrial phenotypes

Author

Listed:
  • Lun Yao

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Kiyan Shabestary

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Sara M. Björk

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Johannes Asplund-Samuelsson

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Haakan N. Joensson

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Michael Jahn

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Elton P. Hudson

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

Abstract

Cyanobacteria are model organisms for photosynthesis and are attractive for biotechnology applications. To aid investigation of genotype-phenotype relationships in cyanobacteria, we develop an inducible CRISPRi gene repression library in Synechocystis sp. PCC 6803, where we aim to target all genes for repression. We track the growth of all library members in multiple conditions and estimate gene fitness. The library reveals several clones with increased growth rates, and these have a common upregulation of genes related to cyclic electron flow. We challenge the library with 0.1 M L-lactate and find that repression of peroxiredoxin bcp2 increases growth rate by 49%. Transforming the library into an L-lactate-secreting Synechocystis strain and sorting top lactate producers enriches clones with sgRNAs targeting nutrient assimilation, central carbon metabolism, and cyclic electron flow. In many examples, productivity can be enhanced by repression of essential genes, which are difficult to access by transposon insertion.

Suggested Citation

  • Lun Yao & Kiyan Shabestary & Sara M. Björk & Johannes Asplund-Samuelsson & Haakan N. Joensson & Michael Jahn & Elton P. Hudson, 2020. "Pooled CRISPRi screening of the cyanobacterium Synechocystis sp PCC 6803 for enhanced industrial phenotypes," Nature Communications, Nature, vol. 11(1), pages 1-13, December.
    • Handle: RePEc:nat:natcom:v:11:y:2020:i:1:d:10.1038_s41467-020-15491-7
    DOI: 10.1038/s41467-020-15491-7
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    Cited by:

    1. Jiao Liu & Moshi Liu & Tuo Shi & Guannan Sun & Ning Gao & Xiaojia Zhao & Xuan Guo & Xiaomeng Ni & Qianqian Yuan & Jinhui Feng & Zhemin Liu & Yanmei Guo & Jiuzhou Chen & Yu Wang & Ping Zheng & Jibin Su, 2022. "CRISPR-assisted rational flux-tuning and arrayed CRISPRi screening of an l-proline exporter for l-proline hyperproduction," Nature Communications, Nature, vol. 13(1), pages 1-16, December.
    2. Yixin Rong & Adrian Frey & Emre Özdemir & Arrate Larrea & Songyuan Li & Alex Toftgaard Nielsen & Sheila Ingemann Jensen, 2024. "CRISPRi-mediated metabolic switch enables concurrent aerobic and synthetic anaerobic fermentations in engineered consortium," Nature Communications, Nature, vol. 15(1), pages 1-8, December.

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