Research progress in carbon neutrality oriented adaptive laboratory evolution of microalgae

doi:10.12211/2096-8280.2021-096

Abstract

Abstract:

Microalgae biotechnology is one of the potential ways to realize carbon peaking and carbon neutrality. At present, microalgae have key problems such as low carbon sequestration efficiency, low photosynthetic transformation efficiency and low content of active components. There are also some technological problems which greatly limit the pace of its industrialization. Most microalgae can not tolerate more than 2% CO₂. Apart from 10%～25% CO₂, there are other pollutants such as NO _x and SO _x in industrial flue gas. These flue gas components inhibit the growth of microalgae. If the tolerance of algal strains are not enhanced, microalgae can not achieve the goal of stable carbon sequestration. In order to solve the problems of microalgae industrialization, wastewater resources can be used to meet the water demand in microalgae cultivation, and the economy can be improved by growing high value-added products. It is necessary to construct new algae strains by means of biotechnology such as synthetic biology, and build a new technical route of carbon reduction or negative carbon according to the characteristics of microalgae carbon sequestration and metabolism. Adaptive laboratory evolution (ALE) has made some progress in improving CO₂ fixation by microalgae, enhancing wastewater treatment and improving metabolic phenotype. Evolved algal strains resistant to high concentration of carbon dioxide and other environmental stresses have been achieved. However, the efficiency of ALE in microalgae needs to be improved, and there are few studies on the mining of synthetic biological elements based on carbon sequestration, photosynthesis and biosynthesis of active components. In order to overcome the above problems, it is urgent to change the strategy of ALE in microalgae, combined with the application of high-throughput ALE device to speed up the evolution process; Based on the existing evolved strains, the elements of tolerance genes, photosynthesis and biosynthesis of active components will be deeply excavated to lay a foundation for microalgae genetic transformation. It is vital for us to learn carefully from the existing experience of ALE in microorganisms, understand throughly the dynamic process of ALE in microalgae, and explore profoundly the basic law of ALE. Finally, the possible ways of laboratory adaptive evolution to meet the challenge of microalgae carbon neutrality are prospected.

Key words: microalgae, adaptive laboratory evolution, carbon neutrality, CO₂ fixation, synthetic biology

摘要：

微藻生物技术是实现碳达峰和碳中和的潜在途径之一。目前，微藻存在固碳效率低、光合转化效率低以及活性组分含量低等关键问题，需要通过合成生物学等生物技术手段构建新的藻株，并依据微藻固碳和代谢的特点，构筑减碳或负碳的新技术路线。适应性实验室进化（ALE）在提高微藻对二氧化碳固定，强化废水处理和改善代谢表型等方面均取得了一定进展，已获得了耐受高浓度二氧化碳和其他环境压力的进化藻株。但是，微藻ALE的效率还有待提高，基于固碳、光合和活性组分生物合成的合成生物学元件挖掘的研究还比较少。为克服以上问题，亟需改变微藻ALE的策略，结合高通量ALE装置的应用，缩短进化时间；在已有进化株的基础上，深入挖掘耐受基因、光合和活性组分生物合成的元件，为微藻基因改造打下基础；借鉴其他微生物ALE的已有经验，深刻理解微藻实验室适应性进化的动态过程，探索ALE的基本规律。最后对ALE应对微藻碳中和挑战的可能途径进行了展望。

关键词: 微藻, 适应性实验室进化, 碳中和, 固碳, 合成生物学

CLC Number:

Q819

Quanyu ZHAO. Research progress in carbon neutrality oriented adaptive laboratory evolution of microalgae[J]. Synthetic Biology Journal, 2022, 3(5): 901-914.

赵权宇. 面向碳中和的微藻适应性实验室进化研究进展[J]. 合成生物学, 2022, 3(5): 901-914.

Figures/Tables 4

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出发藻株	适应进化压力	接种细胞密度	总时间/d	文献
Desmodesmus sp. MAS1, Heterochlorella sp. MAS3	pH 3.0	—	14	[35]
Chlamydomonas reinhardtii	20 μmol/(m²·s), 60 μmol/(m²·s), 400 μmol/(m²·s)	—	—	[36]
Coccomyxa sp.M2	pH 3.0～7.0	—	—	[37]
Chlorella sp. LAMB 3, Chlorella sp. LAMB 122	40% CO₂	0.4 g/L	7	[38]
Chlorellapyrenoidosa	1500～4750 mg/L NH₄⁺	1×10⁸ cells/mL	19	[39]
Scenedesmus sp.	盐浓度0.5%～2.5% 光强4.0～6.0 klux 温度30～35 ℃	0.1 g/L 0.1 g/L 0.1 g/L	10 10 10	[40]
Chlamydomonas reinhardtii	0.2 mol/L NaCl	5×10⁵ cells/mL	2	[41]

出发藻株	适应进化压力	接种细胞密度	总时间/d	文献
Desmodesmus sp. MAS1, Heterochlorella sp. MAS3	pH 3.0	—	14	[35]
Chlamydomonas reinhardtii	20 μmol/(m²·s), 60 μmol/(m²·s), 400 μmol/(m²·s)	—	—	[36]
Coccomyxa sp.M2	pH 3.0～7.0	—	—	[37]
Chlorella sp. LAMB 3, Chlorella sp. LAMB 122	40% CO₂	0.4 g/L	7	[38]
Chlorellapyrenoidosa	1500～4750 mg/L NH₄⁺	1×10⁸ cells/mL	19	[39]
Scenedesmus sp.	盐浓度0.5%～2.5% 光强4.0～6.0 klux 温度30～35 ℃	0.1 g/L 0.1 g/L 0.1 g/L	10 10 10	[40]
Chlamydomonas reinhardtii	0.2 mol/L NaCl	5×10⁵ cells/mL	2	[41]

出发藻株	适应进化压力	接种细胞密度 /(g/L)	周期	总时间/d	文献
Chlorococcum littorale	缺氮	OD₇₅₀=0.5	13	134（前8个周期，每个周期前2天正常，后面6天缺氮；后面5个周期，前2天正常，后面12天缺氮）	[44]
Chlorella vulgaris	255 μE/(m²·s)红光	0.84	38	114	[45]
Dunaliella salina	42 μE/(m²·s)蓝光 + 128 μE/(m²·s)红光	0.5	16	80	[46]
Dunaliella salina	42 μE/(m²·s)蓝光 + 128 μE/(m²·s)红光 + 2.5 mol/L NaCl	—	—	—	[47]
Tisochrysis lutea	温度波动（低温6～21 ℃, 高温26～35 ℃）	—	13	150	[48]
Synechocystis sp. PCC 6803	3% NaCl	OD₇₃₀=0.2	43	303	[49]
Schizochytrium sp.	34.5 ℃	—	70	70	[49]
Chlamydomonas sp.	5%或7%海盐	0.02	—	84	[50]
Isochrysis galbana Parke	100 mg/L或200 mg/L苯酚	5×10⁵ cells/mL	30	90	[51]
Crypthecodiniumcohnii	9～54 g/L葡萄糖	OD₄₇₀=0.1	260	650	[52]
藻菌共培养	体积分数20%渗滤液	—	21天一个周期	24个月	[53]
C．reinhardtii	0.2 mol/L NaCl	5×10⁵ cells/mL	2～3天一个周期	17个月	[41]
Schizochytrium sp.	30 g/L NaCl	—	150	150	[54]
Schizochytrium sp.	4 ℃, 30 g/L NaCl	—	30	90	[55]
Schizochytrium sp.	230 r/min	—	40	40	[56]
Phaeodactylumtricornutum	50%～90%盐度	1×10⁶ cells/mL	35	252	[32]

出发藻株	适应进化压力	接种细胞密度 /(g/L)	周期	总时间/d	文献
Chlorococcum littorale	缺氮	OD₇₅₀=0.5	13	134（前8个周期，每个周期前2天正常，后面6天缺氮；后面5个周期，前2天正常，后面12天缺氮）	[44]
Chlorella vulgaris	255 μE/(m²·s)红光	0.84	38	114	[45]
Dunaliella salina	42 μE/(m²·s)蓝光 + 128 μE/(m²·s)红光	0.5	16	80	[46]
Dunaliella salina	42 μE/(m²·s)蓝光 + 128 μE/(m²·s)红光 + 2.5 mol/L NaCl	—	—	—	[47]
Tisochrysis lutea	温度波动（低温6～21 ℃, 高温26～35 ℃）	—	13	150	[48]
Synechocystis sp. PCC 6803	3% NaCl	OD₇₃₀=0.2	43	303	[49]
Schizochytrium sp.	34.5 ℃	—	70	70	[49]
Chlamydomonas sp.	5%或7%海盐	0.02	—	84	[50]
Isochrysis galbana Parke	100 mg/L或200 mg/L苯酚	5×10⁵ cells/mL	30	90	[51]
Crypthecodiniumcohnii	9～54 g/L葡萄糖	OD₄₇₀=0.1	260	650	[52]
藻菌共培养	体积分数20%渗滤液	—	21天一个周期	24个月	[53]
C．reinhardtii	0.2 mol/L NaCl	5×10⁵ cells/mL	2～3天一个周期	17个月	[41]
Schizochytrium sp.	30 g/L NaCl	—	150	150	[54]
Schizochytrium sp.	4 ℃, 30 g/L NaCl	—	30	90	[55]
Schizochytrium sp.	230 r/min	—	40	40	[56]
Phaeodactylumtricornutum	50%～90%盐度	1×10⁶ cells/mL	35	252	[32]

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