集胞藻酰基载体蛋白基因过量表达对脂肪酸合成的影响

doi:10.3969/j.issn.1004-1524.2020.07.14

浙江农业学报 ›› 2020, Vol. 32 ›› Issue (7): 1253-1262.DOI: 10.3969/j.issn.1004-1524.2020.07.14

集胞藻酰基载体蛋白基因过量表达对脂肪酸合成的影响

路晓媛¹, 钟怀荣², 夏志洁¹, 曹月蕾¹, 陈高^2,*, 戴美学^1,*

1.山东师范大学生命科学学院,山东济南 250014;
2.山东省农业科学院生物技术研究中心,山东济南 250100

收稿日期:2019-12-02 出版日期:2020-07-25 发布日期:2020-07-28
通讯作者: *陈高,E-mail:gxchen001@hotmail.com;戴美学,E-mail: daimeixue@sdnu.edu.cn
作者简介:路晓媛(1995—),女,山东烟台人,硕士研究生,主要从事藻类分子生物学研究。E-mail:luxiaoyuan1225@163.com
基金资助:
山东省重点研发计划(公益性科技攻关类)(2018GSF121019,2019GSF107098); 2019年度中国科学院藻类生物学重点实验室开放课题(2018-006); 山东省科技发展计划(2013GSF12116); 国家重点研发计划政府间国际科技创新合作/港澳台科技创新合作重点专项(2018YFE0108600)

Effects of overexpression of acyl carrier protein gene in Synechocystis on fatty acids synthesis

LU Xiaoyuan¹, ZHONG Huairong², XIA Zhijie¹, CAO Yuelei¹, CHEN Gao^2,*, DAI Meixue^1,*

1. School of Life Science, Shandong Normal University, Jinan 250014, China;
2. Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Jinan 250100, China

Received:2019-12-02 Online:2020-07-25 Published:2020-07-28

摘要/Abstract

摘要： 酰基载体蛋白(ACP)是微生物脂肪酸生物合成途径中的一个关键蛋白。为了促进集胞藻脂肪酸的积累,以集胞藻PCC6803为研究对象,成功构建了用于过量表达ACP(ssl2084)基因的同源重组质粒ssl2084⁽⁺⁾并在集胞藻PCC6803中进行转化。将ssl2084⁽⁺⁾突变株进行DNA水平和蛋白质水平鉴定,结果表明,ssl2084基因已经得到过量表达。利用气相色谱—质谱联用技术(GC-MS)对突变藻株在不同温度下脂肪酸的含量及组分进行检测,发现在30 ℃培养条件下ssl2084⁽⁺⁾突变株中C12:0、C16:0和C18:0的含量显著增加,分别为1.50、8.74和0.60 mg·g^-1,与野生型相比分别增加了54.71%、50.82%和155.86%;在20 ℃低温胁迫培养条件下,C12:0、C16:0和C18:0的含量分别达到1.70、11.85和2.31 mg·g^-1,与野生型相比分别增加了31.94%、47.35%和39.90%。以上结果表明,在集胞藻PCC6803中过表达ssl2084基因能提高集胞藻中的中长链脂肪酸的含量,低温条件使得集胞藻中的中长链脂肪酸的含量进一步提高。

关键词: 集胞藻PCC6803, 酰基载体蛋白, 同源重组, 中长链脂肪酸

Abstract: Acyl carrier protein (ACP) is a key protein in fatty acid biosynthesis pathway of microorganisms. In order to promote the accumulation of fatty acids in Synechocystis, the homologous recombinant plasmid ssl2084⁽⁺⁾ was successfully constructed for overexpressing the ACP (ssl2084) gene and transformed into wild-type Synechocystis sp. PCC6803. Identification of the ssl2084⁽⁺⁾ mutant strain at the DNA and protein levels showed that the ssl2084 gene had been overexpressed. The content and composition of the fatty acids in the mutant strain under different temperatures were detected by liquid chromatography-mass spectrometry (GC-MS). The results showed that the contents of C12:0, C16:0 and C18:0 in ssl2084⁽⁺⁾ mutant at 30 ℃ were 1.50, 8.74 and 0.60 mg·g^-1, respectively, which increased by 54.71%, 50.82% and 155.86%, respectively, compared with the wild type. Under the condition of low temperature stress culture at 20 ℃, the contents of C12:0, C16:0 and C18:0 reached 1.70, 11.85 and 2.31 mg·g^-1, respectively, which increased by 31.94%, 47.35% and 39.90%, respectively, compared with the wild type. The above results indicated that overexpression of ssl2084 gene in Synechocystis sp. PCC6803 could increase the content of medium and long chain fatty acids, and the low temperature condition could further increase the content of medium and long chain fatty acids.

Key words: Synechocystis sp. PCC6803, acyl carrier protein, homologous recombination, medium and long chain fatty acid

中图分类号:

Q786

路晓媛, 钟怀荣, 夏志洁, 曹月蕾, 陈高, 戴美学. 集胞藻酰基载体蛋白基因过量表达对脂肪酸合成的影响[J]. 浙江农业学报, 2020, 32(7): 1253-1262.

LU Xiaoyuan, ZHONG Huairong, XIA Zhijie, CAO Yuelei, CHEN Gao, DAI Meixue. Effects of overexpression of acyl carrier protein gene in Synechocystis on fatty acids synthesis[J]. , 2020, 32(7): 1253-1262.

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集胞藻酰基载体蛋白基因过量表达对脂肪酸合成的影响

Effects of overexpression of acyl carrier protein gene in Synechocystis on fatty acids synthesis

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