《植物生理学报》 2021, 57(1): 101-111

异源表达猫爪藤ACP-Δ⁹脱氢酶基因促进莱茵衣藻油脂合成

宋亚楠¹，岳敏¹，石颖¹，朱琴¹，薛金爱¹，黄观钦²，季春丽¹，张春辉¹，崔红利^1,*，李润植^1,*

¹山西农业大学分子农业与生物能源研究所, 山西晋中030801；²深圳大学生命与海洋科学学院, 广东深圳518000

通信作者：崔红利；E-mail: cuihongli@sxau.edu.cn；李润植；E-mail: rli2001@126.com

摘　要：

棕榈油酸(C16:1^Δ9)等ω-7脂肪酸是一类单不饱和脂肪酸, 具有重要的营养、医药和工业价值。猫爪藤(Dolichandra unguis-cati)种子富含棕榈油酸, 种子中的酰基-ACP-^Δ9 脱氢酶(DuACP-Δ9D)能高效催化棕榈酸(C16:0)生成棕榈油酸。为获得富含ω-7脂肪酸的优异藻株, 将DuACP-Δ9D基因组成型表达载体pCAMBIA1303-DuACP-Δ9D通过玻璃珠法导入莱茵衣藻(Chlamydomonas reinhardtii) CC849藻细胞, Du-ACP-Δ9D基因在藻细胞中成功表达。与野生型莱茵衣藻CC849相比, 转化藻株1和2中总油脂积累显著增加, 尤其是棕榈酸含量分别提高到8.64和8.22 mg·g⁻¹ 。转化藻株的光合作用和生长特性未现负效应。硫胁迫处理导致转化藻株的总脂和棕榈油酸含量进一步显著上升。转化藻株碳水化合物和蛋白质含量则明显降低。总之, 异源表达DuACP-Δ9D基因可显著提高微藻细胞总油脂积累和ω-7脂肪酸的富集, 且未对其他性状产生不利影响。研究为未来应用微藻规模化生产高附加值ω-7油脂提供新种质。

关键词：猫爪藤DuACP-Δ9D基因; 莱茵衣藻; 油脂; ω-7脂肪酸; 异源表达

收稿：2020-07-29   修定：2020-11-27

资助：国家自然科学基金(31902394)、山西省应用基础研究计划项目(201801D22125)、山西省重点研发计划项目(201803D31063和 201603D312005)、山西农谷建设科研专项(SXNGJSKYZX201906)、晋中市重点研发计划项目(Y192012)、山西农业大学科技创 新基金(2018YJ16)、山西省煤基重大科技专项(FT-2014-01)、山西省研究生创新项目(2020BY056)和山西省留学基金(2015-064)

Heterologous expression of Dolichandra unguis-cati ACP Δ⁹ dehydrogenase gene promotes lipid synthesis in Chlamydomonas reinhardtii

SONG Yanan¹, YUE Min¹, SHI Ying¹, ZHU Qin¹, XUE Jinai¹, HUANG Guanqin², JI Chunli¹, ZHANG Chunhui¹, CUI Hongli^1,*, LI Runzhi^1,*

¹Institute of Molecular Agriculture & Bioenergy, Shanxi Agricultural University, Jinzhong, Shanxi 030801, China; ²College of Life Sciences and Oceanography, Shenzhen University, Shenzhen, Guangdong 518000, China

Corresponding author: CUI Hongli; E-mail: cuihongli@sxau.edu.cn; LI Runzhi; E-mail: rli2001@126.com

Abstract:

Ω-7 fatty acids such as palmitoleic acid (C16:1^Δ9 ) are a group of monounsaturated fatty acids

with important application value for human nutrition, pharmaceuticals, and industries. Dolichandra un-

guis-cati seed oil is rich in palmitoleic acid. An acyl-ACP Δ⁹ desaturase (DuACP-Δ9D) derived from developing seeds of this species can effectively catalyze the conversion of palmitic acid (C16:0) to palmitoleic acid. In order to develop excellent algal strains rich in ω-7 fatty acids, a constitutive expression vector (pCAMBIA1303-DuACP-Δ9D) bearing DuACP-Δ9D gene was transformed into Chlamydomonas reinhardtii CC849 by glass bead method. The results demonstrate that DuACP-Δ9D gene was expressed successfully in the algal cells. Compared to wild-type C. reinhardtii CC849, the transgenic algal strains 1 and 2 significantly increased total lipid and palmitoleic acid accumulation, particularly levels of palmitoleic acid up to 8.64 and 8.22 mg·g⁻¹ , respectively. No negative effect was detected in these transgenic algal strains. Moreover, sulfur stress resulted in further significant enhancement of total lipid and palmitoleic acid contents in the transgenic algae. Meanwhile, carbohydrate and protein contents in the transgenic strains were largely decreased under sulfur stress. Taken together, heterologous overexpression of DuACP-Δ9D gene can significantly increase the accumulation of total lipid and ω-7 fatty acids in microalgal cells without adverse impact on other straits. The present study provides new algal stains for commercial production of high-valued oils enriched with ω-7 fatty acids in the future.

Key words: Dolichandra unguis-cati DuACP-Δ9D gene; Chlamydomonas reinhardtii; lipid; ω-7 fatty acid; het- erologous expression