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Development of a primary culture system for haematopoietic tissue cells from Cherax quadricarinatus and an exploration of transfection methods
Xu, Xiaohui1,2; Duan, Hu1,2; Shi, Yingli1,2; Xie, Shijun1,2; Song, Zhan3; Jin, Songjun1,2; Li, Fuhua1,2; Xiang, Jianhai1,2
2018-11-01
Source PublicationDEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY
ISSN0145-305X
Volume88Pages:45-54
Corresponding AuthorXiang, Jianhai(jhxiang@qdio.ac.cn)
AbstractVarious known and unknown viral diseases can threaten crustacean aquaculture. To develop prophylactic and therapeutic strategies against viruses, crustacean cell lines are urgently needed for immunology and virology studies. However, there are currently no permanent crustacean cell lines available. In this study, we developed a new method for preparing crayfish plasma (CP) and found that CP enhanced the proliferative capacity of haematopoietic tissue (hpt) cells from Cherax quadricarinatus by an EdU (5-ethynyl-2'-deoxyuridine) assay. The optimal CP concentration for hpt cell culture and the optimal subculture method are discussed. To achieve efficient expression of a foreign gene in hpt cells cultured in vitro, different transfection methods and vectors were analysed. We found that Lipofectamine 2000 could be used to efficiently transfect a foreign vector into hpt cells and exhibited a lower level of cytotoxicity than the other methods tested, and transfection of pEGFP-N1/w249 and pDHsp70-EGFP-FLAG resulted in high EGFP expression. By transmission electron microscopy (TEM) and virus copy number analysis, we found that white spot syndrome virus (WSSV) could infect hpt cells and multiply efficiently. Our results implied that the crayfish hpt cell culture system we improved could be used as a replacement for immortal crustacean cell lines in viral infection studies. Our findings provide a solid foundation for future immortalization and gene function studies in crustacean cells.
KeywordCherax quadricarinatus Culture of haematopoietic tissue cells Transfection WSSV infection
DOI10.1016/j.dci.2018.07.006
Indexed BySCI
Language英语
Funding ProjectMajor State Basic Research Development Programs of China (973 Program)[2012CB114403] ; Qingdao National Laboratory for Marine Science and Technology[2015ASKJ02]
WOS Research AreaFisheries ; Immunology ; Zoology
WOS SubjectFisheries ; Immunology ; Zoology
WOS IDWOS:000443668600006
PublisherELSEVIER SCI LTD
Citation statistics
Document Type期刊论文
Identifierhttp://ir.qdio.ac.cn/handle/337002/160272
Collection实验海洋生物学重点实验室
Corresponding AuthorXiang, Jianhai
Affiliation1.Chinese Acad Sci, Inst Oceanol, Key Lab Expt Marine Biol, 7 Nanhai Rd, Qingdao 266071, Peoples R China
2.Qingdao Natl Lab Marine Sci & Technol, Lab Marine Biol & Biotechnol, Qingdao, Peoples R China
3.Ocean Univ China, Coll Marine Life Sci, Qingdao, Peoples R China
First Author AffilicationInstitute of Oceanology, Chinese Academy of Sciences
Corresponding Author AffilicationInstitute of Oceanology, Chinese Academy of Sciences
Recommended Citation
GB/T 7714
Xu, Xiaohui,Duan, Hu,Shi, Yingli,et al. Development of a primary culture system for haematopoietic tissue cells from Cherax quadricarinatus and an exploration of transfection methods[J]. DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY,2018,88:45-54.
APA Xu, Xiaohui.,Duan, Hu.,Shi, Yingli.,Xie, Shijun.,Song, Zhan.,...&Xiang, Jianhai.(2018).Development of a primary culture system for haematopoietic tissue cells from Cherax quadricarinatus and an exploration of transfection methods.DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY,88,45-54.
MLA Xu, Xiaohui,et al."Development of a primary culture system for haematopoietic tissue cells from Cherax quadricarinatus and an exploration of transfection methods".DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY 88(2018):45-54.
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