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<p><i>Chlamydomonas reinhardtii</i> CW15, a cell-wall-deficient strain, is a widely used chassis in microalgal genetic engineering. However, its low cell density under autotrophic and mixotrophic cultivation restricts industrial scalability. This study aimed to establish a cost-effective, high-density heterotrophic cultivation process for <i>C. reinhardtii</i> CW15. Initially, an optimized AP medium (Tris-free TAP with K<sub>2</sub>HPO<sub>4</sub> as the sole phosphorus source) significantly reduced medium costs while achieving a cell density of 0.446 g/L. Subsequently, a fed-batch process in a 5 L fermentor yielded a record cell density of 22.1 g/L (growth rate: 94.6 mg/L·h) at 233 h by optimizing nutrient supplementation. Scaling up to a 50 L fermentor revealed challenges in ammonia volatilization and salinity accumulation during sterilization. These were resolved by substituting the nitrogen source with ammonia water and supplementing ammonium acetate as a dual nitrogen and partial carbon source, ultimately achieving 30.2 g/L (140 mg/L·h) after 215 h—the highest reported density for cell-wall-deficient <i>C. reinhardtii</i>. This work provides an efficient and scalable cultivation strategy, facilitating its industrial and biotechnological applications.</p>

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High-density heterotrophic cultivation of a cell-wall-deficient Chlamydomonas reinhardtii strain by fed-batch strategy

  • 期刊论文

  • 发表日期：

    2025年7月3日

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  • Kaidi Zhou,
  • Tao Yu,
  • Minxi Wan,
  • Zhengxu Bao,
  • Wenming Bai,
  • Weiliang Wang,
  • Yuanguang Li

摘要

Chlamydomonas reinhardtii CW15, a cell-wall-deficient strain, is a widely used chassis in microalgal genetic engineering. However, its low cell density under autotrophic and mixotrophic cultivation restricts industrial scalability. This study aimed to establish a cost-effective, high-density heterotrophic cultivation process for C. reinhardtii CW15. Initially, an optimized AP medium (Tris-free TAP with K2HPO4 as the sole phosphorus source) significantly reduced medium costs while achieving a cell density of 0.446 g/L. Subsequently, a fed-batch process in a 5 L fermentor yielded a record cell density of 22.1 g/L (growth rate: 94.6 mg/L·h) at 233 h by optimizing nutrient supplementation. Scaling up to a 50 L fermentor revealed challenges in ammonia volatilization and salinity accumulation during sterilization. These were resolved by substituting the nitrogen source with ammonia water and supplementing ammonium acetate as a dual nitrogen and partial carbon source, ultimately achieving 30.2 g/L (140 mg/L·h) after 215 h—the highest reported density for cell-wall-deficient C. reinhardtii. This work provides an efficient and scalable cultivation strategy, facilitating its industrial and biotechnological applications.