小球藻的基因工程改造研究进展

doi:10.14012/j.cnki.fjsc.2023.06.010

摘要/Abstract

摘要：

小球藻是一种可以利用阳光和二氧化碳进行光合生长并积累生物质的单细胞微藻,被广泛用于生产生物燃料和其他高附加值产品,在水产养殖、食品、能源和环境等领域均有重要应用。随着基因工程技术的发展,小球藻分子水平方面的研究和应用逐渐受到关注,对其进行特定的遗传改造也成为了可能。本文通过从转化系统、转化方法、转化影响因素、外源蛋白表达等四个方面进行总结归纳,阐述了小球藻在基因工程改造领域中的应用和仍存在的难题,期望为小球藻的高附加值产品生产提供理论支持和参考。

关键词: 小球藻, 基因工程, 转化, 蛋白表达

Abstract:

Chlorella is a unicellular microalga that could grow and accumulate biomass through photosynthesis using sunlight and carbon dioxide. It is widely used to produce biofuels and other high-value products in fields like aquaculture, food, energy, and the environment. With the development of genetic engineering technology, research and applications of Chlorella at the molecular level have received increasing attention, making specific genetic modification possible. In this paper, we summarized and discussed the application of Chlorella in genetic engineering and protein production from four perspectives: transformation system, transformation methods, factors affecting transformation and expression of exogenous proteins. The aim of this review was to provide theoretical support and references for high-value products production of Chlorella.

Key words: Chlorella, genetic engineering, transformation, protein express

中图分类号:

Q939.9

张甜甜, 陈必链, 郑明敏. 小球藻的基因工程改造研究进展[J]. 渔业研究, 2023, 45(6): 593-602.

ZHANG Tiantian, CHEN Bilian, ZHENG Mingmin. Advances of genetic engineering modification in Chlorella[J]. Journal of Fisheries Research, 2023, 45(6): 593-602.

图/表 3

参考文献 56

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小球藻藻种 Chlorella species	抗性基因 Resistant genes	抗生素 Antibiotic	最佳筛选浓度/mg·mL^-1 Optimal screening concentration	参考文献 Reference
普通小球藻C.vulgaris	aadA	奇霉素	0.10~0.20	[6]
椭圆小球藻C.ellipsoidea	NptⅡ	G418	0.03	[36]
蛋白核小球藻C.pyrenoidosa	aadA	硫酸链霉素	0.10	[27]
蛋白核小球藻C.pyrenoidosa		卡那霉素	1.00	[29]
小球藻Chlorella	Kanr	G418	1.00	[20]
椭圆小球藻C.ellipsoidea	Sh ble	腐草霉素	0.001	[40]
普通小球藻C.vulgaris	hpt	潮霉素	0.02	[41]
小球藻Chlorella sp.	hpt	潮霉素B	0.05	[21]
普通小球藻C.vulgaris	hpt	潮霉素B	0.30	[42]
蛋白核小球藻C.pyrenoidosa	NptⅡ	G418	0.03	[11]
普通小球藻C.vulgaris	NptⅡ	G418	0.03	[5]
椭圆小球藻C.ellipsoidea	NptⅡ、NR	G418	0.03	[43]
椭圆小球藻C.ellipsoidea	ble	博来霉素	0.01	[44]
普通小球藻C.vulgaris	CAT	氯霉素	0.20	[45]
普通小球藻C.vulgaris	hpt	潮霉素B	0.02	[33]
小球藻Chlorella	neo	G418	0.16	[23]

小球藻藻种 Chlorella species	抗性基因 Resistant genes	抗生素 Antibiotic	最佳筛选浓度/mg·mL^-1 Optimal screening concentration	参考文献 Reference
普通小球藻C.vulgaris	aadA	奇霉素	0.10~0.20	[6]
椭圆小球藻C.ellipsoidea	NptⅡ	G418	0.03	[36]
蛋白核小球藻C.pyrenoidosa	aadA	硫酸链霉素	0.10	[27]
蛋白核小球藻C.pyrenoidosa		卡那霉素	1.00	[29]
小球藻Chlorella	Kanr	G418	1.00	[20]
椭圆小球藻C.ellipsoidea	Sh ble	腐草霉素	0.001	[40]
普通小球藻C.vulgaris	hpt	潮霉素	0.02	[41]
小球藻Chlorella sp.	hpt	潮霉素B	0.05	[21]
普通小球藻C.vulgaris	hpt	潮霉素B	0.30	[42]
蛋白核小球藻C.pyrenoidosa	NptⅡ	G418	0.03	[11]
普通小球藻C.vulgaris	NptⅡ	G418	0.03	[5]
椭圆小球藻C.ellipsoidea	NptⅡ、NR	G418	0.03	[43]
椭圆小球藻C.ellipsoidea	ble	博来霉素	0.01	[44]
普通小球藻C.vulgaris	CAT	氯霉素	0.20	[45]
普通小球藻C.vulgaris	hpt	潮霉素B	0.02	[33]
小球藻Chlorella	neo	G418	0.16	[23]

小球藻藻种 Chlorella species	表达载体 Vector	启动子 Promoter	转化方法 Transformation method	转化率或蛋白表达量 Transformation rate or protein expression	参考文献 Reference
普通小球藻 C.vulgaris	pUC改造的 pCMCC	Prrn	电穿孔	人碱性成纤维细胞生长因子表达量0.26~1.42 ng/g藻细胞鲜重	[37]
普通小球藻 C.vulgaris	pMD18T	P16S、rbcL	微粒子轰击	转化率(212±48)转化子/μg质粒DNA	[6]
索罗金小球藻 C.sorokiniana	pCAMBIA1301	CaMV35S	农杆菌介导	转化率(220±5)转化子/10⁶细胞	[31]
普通小球藻 C.vulgaris	pBYR2e	CaMV35S	农杆菌介导	表达量1.14 μg/g鲜重、成纤维细胞生长因子表达量1.61 ng/g鲜重	[53]
椭圆小球藻 C.ellipsoidea	pGREEN0029、 pBAC818	e35s	纳米载体介导	转化率(0.95×10²±0.07)转化子/μg质粒DNA	[36]
佐夫色绿藻 C.zofingiensis	pBI121	CaMV35S	农杆菌介导	虾青素质量比 0.564 mg/g	[32]
蛋白核小球藻 C.pyrenoidosa	pBI121		微粒子轰击	目的蛋白含量高达55.94%	[29]
小球藻 Chlorella sp.	pMDC85	2×CaMV35S	电穿孔	可溶性总蛋白中目的蛋白含量约占0.052%~0.078%	[54]
蛋白核小球藻 C.pyrenoidosa	pGREENII0029	Ubi	电穿孔	转化率(101±7)转化子/μg质粒DNA	[11]
普通小球藻 C.vulgaris	pBI121	CaMV35S	PEG介导	转化率(356±30)转化子/μg质粒DNA	[5]
椭圆小球藻 C.ellipsoidea	pGREEN0029	Ubi	电穿孔	转化率750转化子/μg质粒DNA,蛋白表达量11.42 mg/L	[43]
椭圆小球藻 C.ellipsoidea	pSP124	Ubi-1、rbcS2	纤维素酶处理	转化率2.25×10³转化子/μg质粒DNA	[44]

小球藻藻种 Chlorella species	表达载体 Vector	启动子 Promoter	转化方法 Transformation method	转化率或蛋白表达量 Transformation rate or protein expression	参考文献 Reference
普通小球藻 C.vulgaris	pUC改造的 pCMCC	Prrn	电穿孔	人碱性成纤维细胞生长因子表达量0.26~1.42 ng/g藻细胞鲜重	[37]
普通小球藻 C.vulgaris	pMD18T	P16S、rbcL	微粒子轰击	转化率(212±48)转化子/μg质粒DNA	[6]
索罗金小球藻 C.sorokiniana	pCAMBIA1301	CaMV35S	农杆菌介导	转化率(220±5)转化子/10⁶细胞	[31]
普通小球藻 C.vulgaris	pBYR2e	CaMV35S	农杆菌介导	表达量1.14 μg/g鲜重、成纤维细胞生长因子表达量1.61 ng/g鲜重	[53]
椭圆小球藻 C.ellipsoidea	pGREEN0029、 pBAC818	e35s	纳米载体介导	转化率(0.95×10²±0.07)转化子/μg质粒DNA	[36]
佐夫色绿藻 C.zofingiensis	pBI121	CaMV35S	农杆菌介导	虾青素质量比 0.564 mg/g	[32]
蛋白核小球藻 C.pyrenoidosa	pBI121		微粒子轰击	目的蛋白含量高达55.94%	[29]
小球藻 Chlorella sp.	pMDC85	2×CaMV35S	电穿孔	可溶性总蛋白中目的蛋白含量约占0.052%~0.078%	[54]
蛋白核小球藻 C.pyrenoidosa	pGREENII0029	Ubi	电穿孔	转化率(101±7)转化子/μg质粒DNA	[11]
普通小球藻 C.vulgaris	pBI121	CaMV35S	PEG介导	转化率(356±30)转化子/μg质粒DNA	[5]
椭圆小球藻 C.ellipsoidea	pGREEN0029	Ubi	电穿孔	转化率750转化子/μg质粒DNA,蛋白表达量11.42 mg/L	[43]
椭圆小球藻 C.ellipsoidea	pSP124	Ubi-1、rbcS2	纤维素酶处理	转化率2.25×10³转化子/μg质粒DNA	[44]