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bioRxiv · 10.1101/2023.06.30.547221

Biodecomposing Spirulina platensis by a de novo designed Bacillus subtilis-based method to develop a medium for the high cell-density cultivation of Escherichia coli in batch mode

Li, X.·Wei, Z.·Ge, J.·Pan, Y.·Chu, X.·Huang, B.·Zhao, J.·Li, Y.·Zhou, Y.
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Abstract

High cell-density cultivation (HCDC) is fundamental to basic research and industrial applications, especially in batch mode. However, limited media are available for batch culture of HCDC, because the media for batch culture must contain extremely sufficient nutrients on the one hand and few or even no substrates to generate detrimental metabolite on the other hand to attain HCDC. Spirulina platensis (SP), a new media material, is considered ideal for the development of such media. Here, we develop a biolysis method for SP degradation based on the cultivation supernatant of Bacillus subtilis and extensively demonstrate its higher degradation (indicated by the production of more small peptides and free amino acids) and cultivation effectiveness with three other methods. Based on its SP hydrolysates, a modified SP-based broth (MSP) is then formulated. Tests on Escherichia coli K-12 show that MSP achieves HCDC with several benefits: (i) its maximum optical density at 600 nm is [~]16.67, significantly higher than that of Luria-Bertani (LB) broth (only [~]6.30); (ii) MSP requires only 36 h to reach peak growth, much faster than that of LB (48 h); (iii) its maximum growth (1.12 {+/-} 0.01 h-1) is significantly higher than that of LB (0.20 {+/-} 0.00 h-1); (v) MSP initiates growth immediately after inoculation (lag time <0), comparable to LB; (iv) the number of viable cells in MSP is high ([~]2.16 x 1011 ml-1), [~]10.19 times the amount in LB. Consequently, we envision MSP will become the first choice for E. coli HCDC batch culture in the future. ImportanceSo far, it is the first time to develop a high-efficiency method for transforming Spirulina platensis (SP) into medium ingredients. Based on its SP hydrolysates, a high cell-density cultivation (HCDC) medium for the batch culture of Escherichia coli is formulated for the first time, which is greatly beneficial for both basic research and industrial applications. In addition to HCDC, the SP hydrolysates can be extended to a wide range of applications, due to their rich nutrient content. Besides, this study demonstrates for the first time that SP is an ideal material to develop HCDC media. Furthermore, this study demonstrates that medium development and modification for batch culture can attain HCDC, without the development of new culture technologies. Therefore, this study highlights the importance of the rebirth of medium development and modification and supports the shift from developing new culture technologies to medium development and modification for HCDC in batch mode.

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Li, X., Wei, Z., Ge, J., Pan, Y., Chu, X., Huang, B., Zhao, J., Li, Y., Zhou, Y.. 2023-06-30. Biodecomposing Spirulina platensis by a de novo designed Bacillus subtilis-based method to develop a medium for the high cell-density cultivation of Escherichia coli in batch mode. https://doi.org/10.1101/2023.06.30.547221

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