产微球茎属（Microbulbifer sp.）FG4菌株琼胶酶功能基因的克隆与表达

doi:10.3969/j.issn.1001-1803.2023.03.006

日用化学工业（中英文） ›› 2023, Vol. 53 ›› Issue (3): 285-291.doi: 10.3969/j.issn.1001-1803.2023.03.006

产微球茎属（Microbulbifer sp.）FG4菌株琼胶酶功能基因的克隆与表达

张力雄,刘明明,陈良,方再光^*()

海南大学海洋学院，海南海口 570228

收稿日期:2022-03-31 修回日期:2023-02-28 出版日期:2023-03-22 发布日期:2023-03-22
基金资助:
国家自然科学基金资助项目(40666001)

Cloning and expression of a novel gene encoding agarase from Microbulbifer sp. FG4

Zhang Lixiong,Liu Mingming,Chen Liang,Fang Zaiguang^*()

Ocean College, Hainan University, Haikou, Hainan 570228, China

Received:2022-03-31 Revised:2023-02-28 Online:2023-03-22 Published:2023-03-22
Contact: * Tel.: +86-15027423243, E-mail: zl_xiong91@163.com.

摘要/Abstract

摘要：

以FG4菌株琼胶酶基因侧翼序列为基础，构建重组质粒与载体，从而获得高效产琼胶酶的表达菌株。本研究利用染色体步移法获得FG4菌珠琼胶酶基因功能的侧翼序列之后，将基因组端序列克隆基因片段与表达载体进行重组，并转入到受体细胞BL21中，在鉴定表达菌株的成功构建后利用IPTG诱导表达琼胶酶。实现产微球茎属（Microbulbifer sp.） FG4菌株琼胶酶功能基因的克隆与表达，以期为琼胶酶类在不同领域的实际应用提供理论支持。

关键词: 产微球茎属, 琼胶酶, 染色体步移, 克隆与表达

Abstract:

Based on the flanking sequence of the agarase gene of FG4 strain, the recombinant plasmid and vector were constructed to obtain the engineering strain which could express the agarase efficiently. In this study, the flanking sequence of FG4 agarase gene is obtained by chromosome walking. The cloned gene is recombined with the expression vector pET-22b （+）and transferred into the recipient BL21. After the successful construction of the expressed strain, the expression of agarase is induced by IPTG and the optimal fermentation conditions of the expressed strain are explored. This strain ensures the catalytic activity of agarase and lays the foundation for the agarase-based pharmaceutical industry, daily chemical industry and food industry. The cloning and expression of agarase gene from Microbulbifers sp. FG4 strain of Microbulbifers provides theoretical support for agarase application in different fields.

Key words: Micrcbulbifer sp., agarase, chromosome walking, cloning and expression

中图分类号:

TQ658

张力雄, 刘明明, 陈良, 方再光. 产微球茎属（Microbulbifer sp.）FG4菌株琼胶酶功能基因的克隆与表达[J]. 日用化学工业（中英文）, 2023, 53(3): 285-291.

Zhang Lixiong, Liu Mingming, Chen Liang, Fang Zaiguang. Cloning and expression of a novel gene encoding agarase from Microbulbifer sp. FG4[J]. China Surfactant Detergent & Cosmetics, 2023, 53(3): 285-291.

图/表 12

表1

图1

图2

图3

图4

图5

图6

图7

图8

图9

图10

图11

参考文献 13

[1]	Wu H, Wang K, Wang H, et al. Novel self-assembled tacrolimus nanoparticles cross-linking thermosensitive hydrogels for local rheumatoid arthritis therapy[J]. Colloids and Surfaces B: Biointerfaces, 2017, 149: 97-104. doi: S0927-7765(16)30717-2 pmid: 27736727
[2]	Arnott S, Fulmer A, Scott W E, et al. The agarose double helix and its function in agarose gel structure[J]. J. Mol. Biol., 1974, 90 (2) : 269-284. pmid: 4453017
[3]	Li Rong, Liang Jiaming, Li Haixin, et al. Orthogonal experimental study on agar oligosaccharide produced by agarase hydrolysis of agar[J]. Guangdong Feed, 2018, 27 (7) : 43-45.
[4]	Bianco F, Monteverde G, Race M, et al. Comparing performances, costs and energy balance of ex situ remediation processes for PAH-contaminated marine sediments[J]. Environmental Science and Pollution Research, 2020, 27 (16) : 19363-19374. doi: 10.1007/s11356-020-08379-y
[5]	Pramanik K, Mandal S, Banerjee S, et al. Unraveling the heavy metal resistance and biocontrol potential of Pseudomonas sp. K32 strain facilitating rice seedling growth under Cd stress[J]. Chemosphere, 2021, 274 (4) : 129819. doi: 10.1016/j.chemosphere.2021.129819
[6]	Jf Ga B, Tv A, Mu A, et al. Description of three new Alteromonas species Alteromonas antoniana sp. nov. Alteromonas lipotrueae sp. nov. and Alteromonas lipotrueiana sp. nov. isolated from marine environments, and proposal for reclassification of the genus Salinimonas as Alteromonas[J]. Systematic and Applied Microbiology, 2021, 44 (4).
[7]	Park Y, Maeng S, Han J H, et al. Rhodocytophaga rosea sp. nov. and Nibribacter ruber sp. nov. two radiation-resistant bacteria isolated from soil[J]. Antonie van Leeuwenhoek, 2020, 113 (12) : 1-9. doi: 10.1007/s10482-019-01354-9
[8]	Jurnal B, Lingkungan B, Industri K, et al. Evaluation on methods for identification of vibrio sp.: kawakawa (euthynnus affinis) brining shredded and chitosan addition as preservative[J]. Biolink (Jurnal Biologi Lingkungan Industri Kesehatan), 2021, 8 (1) : 103-113. doi: 10.31289/biolink.v8i1
[9]	Cai Y, Wang R, Rao P, et al. Bioremediation of petroleum hydrocarbons using Acinetobacter sp. SCYY-5 isolated from contaminated oil sludge: strategy and effectiveness study[J]. International Journal of Environmental Research and Public Health, 2021, 18 (2) : 819. doi: 10.3390/ijerph18020819
[10]	Wang X, Guo F, Tian P, et al. Flammeovirga agarivorans sp. nov. an agar-digesting marine bacterium isolated from surface seawater[J]. International Journal of Systematic and Evolutionary Microbiology, 2020, 70 (12).
[11]	Li T, Mann R, Kaur J, et al. Transcriptome analyses of barley roots inoculated with novel Paenibacillus sp. and erwinia gerundensis strains reveal beneficial early-stage plant-bacteria interactions[J]. Plants, 2021, 10 (9) : 1802. doi: 10.3390/plants10091802
[12]	Shi X, Guan R, Zhou L, et al. Structural characterization and heparanase inhibitory activity of fucosylated glycosaminoglycan from holothuria floridana[J]. Marine Drugs, 2021, 19 (3) : 162. doi: 10.3390/md19030162
[13]	Pan H B, Wai C M, Kuo L J, et al. A highly efficient uranium grabber derived from acrylic fiber for extracting uranium from seawater[J]. Dalton Transactions, 2020, 49 (9).

试剂	使用量/μL
FG4 DNA	1
dNTP Mixture （2.5 mmol/L each）	8
10×LA PCR Buffer II （Mg²⁺ plus）	5
TaKaRa LA Taq （5 U/μL）	0.5
AP1/AP2/AP3/AP4 Primer （100 pmol/μL）	1
FG4SP1 Primer （10 pmol/μL）	1
dH₂O	33.5

产微球茎属（Microbulbifer sp.）FG4菌株琼胶酶功能基因的克隆与表达

Cloning and expression of a novel gene encoding agarase from Microbulbifer sp. FG4

RichHTML

PDF (PC)

赞

可视化

摘要/Abstract

引用本文

使用本文

图/表 12

参考文献 13

相关文章 1

Metrics

本文评价

推荐阅读 10