论文总字数：18476字

摘 要

二十二碳六烯酸（docosahexaenoic acid，DHA）是人体所必需的一种多不饱和脂肪酸，可以促进婴幼儿视网膜细胞成熟、促进大脑生长发育，还能够预防及治疗各类心血管疾病和老年痴呆，具有非常高的价值。裂殖壶菌因具有合成富含DHA油脂的能力而收到了越来越多的关注。裂殖壶菌是一种海洋真菌，由于生长在浓度较高的海水中，所以对生长环境的渗透压有较高的要求。本实验考察了原始菌株和盐驯化菌株在发酵过程的差异，从生物量、油脂产量、脂肪酸组分等几个方面进行对比，并进行转录组学研究，具体研究中心碳的代谢和脂肪酸的合成。结果表明，驯化菌株的生物量达到了134.5g/L，比原始菌株增加了32.77%；油脂产量高达80.14g/L，比原始菌株高出50.33%。另外，与原始菌株相比，驯化菌株中的ATP柠檬酸裂解酶（ACL）、丙酮酸脱氢酶（PDH）和苹果酸酶（ME）表达分别上调2.86倍，2.02倍和2.57倍；与油脂的生物合成相关的酶如脂肪酸合酶（FAS）、甘油-3-磷酸-O-酰基转移酶（GPAT）和二酰甘油酰基转移酶（DGAT）等酶的相对基因表达分别上调了3.06倍、3.1倍和2.9倍。不难发现，盐驯化菌株在各方面的表现都要优于原始菌株，这也为后期进行菌株优化提供了理论指导。

关键词：裂殖壶菌 渗透压 转录组学

Differential Transcriptomic Study of Schizochytrium sp. in High Salt Stress

Abstract

Docososahexaenoic acid (DHA) is a polyunsaturated fatty acid that is essential for the body. It can promote retinal cell maturation, promote brain growth and. Also, it can prevent and treat cardiovascular diseases and senile dementia. That is to say, DHA plays a very important role. Schizochytrium sp. has received more and more attention due to its ability to synthesize DHA-rich oils. Schizochytrium sp. is a marine fungus with high requirements for the osmotic pressure of the growing environment. This experiment examined the differences in the fermentation process between the original strains and the salt acclimated strains, and compared them in terms of biomass, oil production, fatty acid composition, and conducted transcriptomic studies to specifically study the carbon metabolism and fatty acid synthesis at the center. The results showed that the biomass of the domesticated strains reached 134.5 g/L, an increase of 32.77% compared to the original strain; the oil production was as high as 80.14 g/L, which was 50.33% higher than the original strain. In addition, the expression of ATP citrate lyase (ACL), pyruvate dehydrogenase (PDH) and malic enzyme (ME) in the domesticated strains was up-regulated 2.86-fold, 2.02-fold and 2.57-fold, respectively, compared to the original strain; The relative gene expression of enzymes related to biosynthetic enzymes such as fatty acid synthase (FAS), glycerol-3-phosphate-O-acyltransferase (GPAT) and diacylglycerol acyltransferase (DGAT) was upregulated by 3.06, 3.1 times and 2.9 times.It is not difficult to find that the salt acclimation strains performed better than the original strains in all aspects, which also provided theoretical guidance for the later optimization of strains.

Key Words: Schizochytrium sp.; osmotic pressure ; comparative transcriptomic analysis.

目 录

摘要................................................................................................................................I

Abstract........................................................................................................................II

第一章 文献综述 1

1.1 裂殖壶菌概述 1

1.2 裂殖壶菌的发酵工艺 1

1.3 转录组学研究及其技术 4

1.3.1 转录组学简介 4

1.3.2 转录组学的主要技术 4

1.4 本课题的研究内容及意义 7

1.4.1 本课题的研究内容 7

1.4.2 本课题的研究意义 7

第二章 实验材料与方法 8

2.1 实验材料和仪器 8

2.1.1 菌种 8

2.1.2 主要的试剂与仪器 8

2.1.3 培养基 9

2.2 实验设计 9

2.3 测定方法 10

2.3.1 测定细胞干重和总油脂 10

2.3.2脂肪酸分析 10

2.3.3转录组分析 11

第三章 实验结果与讨论 12

3.1 研究补料分批发酵中驯化菌株的油脂产量和组成 12

3.2 驯化菌株的差异转录组分析 15

3.2.1 中心碳的代谢 15

3.2.2 油脂的生物合成 16

第四章 结论与展望 18

4.1 结论 18

4.2 展望 18

参考文献 19

致谢.............................................................................................................................22

第一章 文献综述

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