{"title":"纳米颗粒影响植物细胞内务基因的表达稳定性。","authors":"Rajendran K Selvakesavan, Gregory Franklin","doi":"10.2147/NSA.S265641","DOIUrl":null,"url":null,"abstract":"Purpose We report on the expression stability of several housekeeping/reference genes that can be used in the normalization of target gene expression in quantitative real-time PCR (qRT-PCR) analysis of plant cells challenged with metal nanoparticles (NPs). Materials and Methods Uniform cell suspension cultures of Hypericum perforatum were treated with 25 mg/l silver and gold NPs (14–15 nm in diameter). Cells were collected after 0.5, 4.0, and 12 h. The total RNA isolated from the cells was analyzed for the stability of ACT2, ACT3, ACT7, EF1-α, GAPDH, H2A, TUB-α, TUB-β, and 18S rRNA genes using qRT-PCR. The cycle threshold (Ct) values of the genes were analyzed using the geNorm, NormFinder, BestKeeper, and RefFinder statistical algorithms to rank gene stability. The stability of the top-ranked genes was validated by normalizing the expression of HYP1. Results The expression of the tested housekeeping genes varied with treatment duration and NP types. EF1-α in gold NP treatment and TUB-α and EF1-α in silver NP treatment ranked among the top three positions. However, none of the genes retained their top ranking with time and across NP types. Conclusion EF1-α can be used as a reference for treatment involving both silver and gold NPs in H. perforatum cells. TUB-α can be used only for silver NP-treated cells. The expression instability of most of the housekeeping genes highlights the importance of systematic standardization of reference genes for NP treatment conditions to draw proper conclusions on the target gene expression.","PeriodicalId":18881,"journal":{"name":"Nanotechnology, Science and Applications","volume":"13 ","pages":"77-88"},"PeriodicalIF":4.9000,"publicationDate":"2020-08-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.2147/NSA.S265641","citationCount":"1","resultStr":"{\"title\":\"Nanoparticles Affect the Expression Stability of Housekeeping Genes in Plant Cells.\",\"authors\":\"Rajendran K Selvakesavan, Gregory Franklin\",\"doi\":\"10.2147/NSA.S265641\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Purpose We report on the expression stability of several housekeeping/reference genes that can be used in the normalization of target gene expression in quantitative real-time PCR (qRT-PCR) analysis of plant cells challenged with metal nanoparticles (NPs). Materials and Methods Uniform cell suspension cultures of Hypericum perforatum were treated with 25 mg/l silver and gold NPs (14–15 nm in diameter). Cells were collected after 0.5, 4.0, and 12 h. The total RNA isolated from the cells was analyzed for the stability of ACT2, ACT3, ACT7, EF1-α, GAPDH, H2A, TUB-α, TUB-β, and 18S rRNA genes using qRT-PCR. The cycle threshold (Ct) values of the genes were analyzed using the geNorm, NormFinder, BestKeeper, and RefFinder statistical algorithms to rank gene stability. The stability of the top-ranked genes was validated by normalizing the expression of HYP1. Results The expression of the tested housekeeping genes varied with treatment duration and NP types. EF1-α in gold NP treatment and TUB-α and EF1-α in silver NP treatment ranked among the top three positions. However, none of the genes retained their top ranking with time and across NP types. Conclusion EF1-α can be used as a reference for treatment involving both silver and gold NPs in H. perforatum cells. TUB-α can be used only for silver NP-treated cells. The expression instability of most of the housekeeping genes highlights the importance of systematic standardization of reference genes for NP treatment conditions to draw proper conclusions on the target gene expression.\",\"PeriodicalId\":18881,\"journal\":{\"name\":\"Nanotechnology, Science and Applications\",\"volume\":\"13 \",\"pages\":\"77-88\"},\"PeriodicalIF\":4.9000,\"publicationDate\":\"2020-08-13\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.2147/NSA.S265641\",\"citationCount\":\"1\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Nanotechnology, Science and Applications\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.2147/NSA.S265641\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2020/1/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q2\",\"JCRName\":\"NANOSCIENCE & NANOTECHNOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Nanotechnology, Science and Applications","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.2147/NSA.S265641","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2020/1/1 0:00:00","PubModel":"eCollection","JCR":"Q2","JCRName":"NANOSCIENCE & NANOTECHNOLOGY","Score":null,"Total":0}
Nanoparticles Affect the Expression Stability of Housekeeping Genes in Plant Cells.
Purpose We report on the expression stability of several housekeeping/reference genes that can be used in the normalization of target gene expression in quantitative real-time PCR (qRT-PCR) analysis of plant cells challenged with metal nanoparticles (NPs). Materials and Methods Uniform cell suspension cultures of Hypericum perforatum were treated with 25 mg/l silver and gold NPs (14–15 nm in diameter). Cells were collected after 0.5, 4.0, and 12 h. The total RNA isolated from the cells was analyzed for the stability of ACT2, ACT3, ACT7, EF1-α, GAPDH, H2A, TUB-α, TUB-β, and 18S rRNA genes using qRT-PCR. The cycle threshold (Ct) values of the genes were analyzed using the geNorm, NormFinder, BestKeeper, and RefFinder statistical algorithms to rank gene stability. The stability of the top-ranked genes was validated by normalizing the expression of HYP1. Results The expression of the tested housekeeping genes varied with treatment duration and NP types. EF1-α in gold NP treatment and TUB-α and EF1-α in silver NP treatment ranked among the top three positions. However, none of the genes retained their top ranking with time and across NP types. Conclusion EF1-α can be used as a reference for treatment involving both silver and gold NPs in H. perforatum cells. TUB-α can be used only for silver NP-treated cells. The expression instability of most of the housekeeping genes highlights the importance of systematic standardization of reference genes for NP treatment conditions to draw proper conclusions on the target gene expression.
期刊介绍:
Nanotechnology, Science and Applications is an international, peer-reviewed, Open Access journal that focuses on the science of nanotechnology in a wide range of industrial and academic applications. The journal is characterized by the rapid reporting of reviews, original research, and application studies across all sectors, including engineering, optics, bio-medicine, cosmetics, textiles, resource sustainability and science. Applied research into nano-materials, particles, nano-structures and fabrication, diagnostics and analytics, drug delivery and toxicology constitute the primary direction of the journal.