Establishment of gene transfer and gene silencing methods in a desiccation-tolerant cell line, Pv11

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• 作者：Yoichiro Sogame ; Jun Okada ; Shingo Kikuta ; Yugo Miyata ; Richard Cornette…
• 关键词：Anhydrobiosis ; RNAi ; Desiccation tolerant cells ; Gene expression system
• 刊名：Extremophiles
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：21
• 期：1
• 页码：65-72
• 全文大小：909KB
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Microbiology; Biotechnology; Biochemistry, general; Microbial Ecology;
• 出版者：Springer Japan
• ISSN：1433-4909
• 卷排序：21

文摘

Larvae of the African midge Polypedilum vanderplanki show extreme desiccation tolerance, known as anhydrobiosis. Recently, the cultured cell line Pv11 was derived from this species; Pv11 cells can be preserved in the dry state for over 6 months and retain their proliferation potential. Here, we attempted to expand the use of Pv11 cells as a model to investigate the mechanisms underlying anhydrobiosis in P. vanderplanki. A newly developed vector comprising a constitutive promoter for the PvGapdh gene allowed the expression of exogenous proteins in Pv11 cells. Using this vector, a stable Pv11 cell line expressing green fluorescence protein (GFP) was established and retained desiccation tolerance. Gene silencing with GFP-specific siRNAs significantly suppressed GFP expression to approximately 7.5–34.6% of that in the non-siRNA-transfected GFP stable line. Establishment of these functional assays will enable Pv11 cells to be utilized as an effective tool to investigate the molecular mechanisms underlying anhydrobiosis.