Establishing primary cell cultures from Branchiostoma belcheri Japanese

Cai, Xiaoqing; Wang, Huamin; Huang, Linxuan; Chen, Juntao; Zhang, Qinfen; Zhang, Yan

doi:10.1007/s11626-013-9579-3

Cited by 1 publication

(1 citation statement)

References 18 publications

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“…Intertidal species are tolerant to a broad range of osmotic pressures, as shown in the successful culture of Crassostrea virginica cells at 425 mOsmol/kg ( 48 ). Studies on other marine invertebrates suggest that marine invertebrate cells can deal with a range of osmolality ( 49 , 50 ) or even lower osmolality than that of their internal fluid ( 51 , 52 ). We show that scallop cells can be cultured at 900–1,100 mOsmol/kg, approximately the osmolality of seawater.…”

Section: Discussionmentioning

confidence: 99%

Gene delivery available in molluscan cells by strong promoter discovered from bivalve-infectious virus

Yoon

Konuma

et al. 2022

Proc. Natl. Acad. Sci. U.S.A.

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Understanding gene functions in marine invertebrates has been limited, largely due to the lack of suitable assay systems. Such a system requires investigative methods that are reproducible and can be quantitatively evaluated, such as a cell line, and a strong promoter that can drive high expression of a transgene. In this study, we established primary cell culture from a marine bivalve mollusc, Mizuhopecten yessoensis . Using scallop primary cells, we optimized electroporation conditions for transfection and carried out a luciferase-based promoter activity assay to identify strong promoter sequences that can drive expression of a gene of interest. We evaluated potential promoter sequences from genes of endogenous and exogenous origin and discovered a strong viral promoter derived from a bivalve-infectious virus, ostreid herpesvirus-1 (OsHV-1). This promoter, we termed OsHV-1 promoter, showed 24.7-fold and 16.1-fold higher activity than the cytomegalovirus immediate early (CMV IE) promoter and the endogenous EF1α promoter, the two most commonly used promoters in bivalves so far. Our GFP assays showed that the OsHV-1 promoter is active not only in scallop cells but also in HEK293 cells and zebrafish embryos. The OsHV-1 promoter practically enables functional analysis of marine molluscan genes, which can contribute to unveiling gene-regulatory networks underlying astonishing regeneration, adaptation, reproduction, and aging in marine invertebrates.

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