Establishment and characterization of novel cell lines derived from six lepidopteran insects collected in the
Establishment and characterization of novel cell lines derived from six lepidopteran insects collected in the
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源自六种鳞翅目昆虫的新型细胞系的建立和表征
DOI：
10.1007/s11626-020-00438-5
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Gaku Akiduki
中科院分区：
文献类型：
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作者：
Kazuyo Watanabe;Isao Kobayashi;Masatsugu Hatakeyama;Takumi Kayukawa;Gaku Akiduki
昆虫细胞系是研究细胞间相互作用和基因功能的重要材料。然而，用于实验的合适的细胞系并不总是可用的，特别是在非模型物种中。本研究以6种鳞翅目昆虫的脂肪体为材料，建立了新的细胞系，它们分别是：黑颈小卷蛾（命名为NARO-Cyku）、红头拟步行虫（NARO-Cehy）、黑点拟步行虫（NARO-Haba）、黄花阔翅夜蛾（NARO-Thol）、舞毒蛾（NARO-Lydi）和美国白蛾（NARO-Hycu）。当由于田间采集而导致样品有限时，幼虫脂肪体是起始物料的理想组织。对于原代培养，将培养基体积保持在最小值以保持脂肪体细胞粘附于烧瓶至关重要。用聚-L-赖氨酸包被烧瓶以有效诱导粘附和细胞分裂。细胞传代50次以上后，用线粒体细胞色素c氧化酶I基因DNA条形码鉴定细胞系的身份。除NARO-Lydi和NARO-Hycu外，所有细胞系均为贴壁细胞，6种细胞系的群体倍增时间范围为1.03至2.49。通过转染表达载体，发现在四种贴壁细胞系中诱导基因表达是可行的，并发现立即早期2和theparxactin 3是有效的基因启动子。结果表明，这些细胞系能够进行基因功能分析。因此，利用我们的方法建立非模式鳞翅目昆虫的细胞系，可以为害虫管理和昆虫利用做出实际贡献。
Insect cell lines are used to study cellular interactions and gene functionsin vitroin several research areas. However, suitable cell lines for experiments are not always available, especially in non-model species. Here, we established novel cell lines derived from fat bodies of six lepidopteran insects:Cydia kurokoi(named NARO-Cyku),Cephonodes hylas(NARO-Cehy),Haritalodes basipunctalis(NARO-Haba),Theretra oldenlandiae(NARO-Thol),Lymantria dispar(NARO-Lydi), andHyphantria cunea(NARO-Hycu) collected in the field. The larval fat body was a promising tissue for the starting material when samples were limited due to field collection. It was critical that the medium volume was kept to a minimum for primary culture to maintain adherence of the fat body cells to the flask. The flask was coated with poly-L-lysine for effective induction of adherence and cell division. The identities of cell lines were confirmed using DNA barcoding with themitochondrial cytochrome c oxidase Igene after cultures were passaged over 50 times. All lines except for NARO-Lydi and NARO-Hycu are adherent cells, and population doubling time of six cell lines ranged from 1.03 to 2.49. Induction of gene expression was practicable in the four adherent cell lines as revealed by transfection of expression vectors and found the immediate early 2 and theBombyx actin3 were effective gene promoters. The results suggest that these cell lines are capable of gene functional analysis. Thus, establishments of cell line using our methods for non-model lepidopterans could make a practical contribution to pest management and insect utilization.