Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines
Summary: The interaction manner and biological function of Rab7 and its effector, Rab-interacting lysosomal protein (RILP), remain unclear in invertebrates. We provide a protocol for detecting the effects of Rab7 and RILP terminals on lysosome and autophagy in Spodoptera frugiperda Sf9 cells with ov...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-12-01
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| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166723006135 |
| _version_ | 1797656129269923840 |
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| author | Gaofeng Cui Zhiyan Jiang Guohua Zhong |
| author_facet | Gaofeng Cui Zhiyan Jiang Guohua Zhong |
| author_sort | Gaofeng Cui |
| collection | DOAJ |
| description | Summary: The interaction manner and biological function of Rab7 and its effector, Rab-interacting lysosomal protein (RILP), remain unclear in invertebrates. We provide a protocol for detecting the effects of Rab7 and RILP terminals on lysosome and autophagy in Spodoptera frugiperda Sf9 cells with overexpression and RNA interference. We describe steps for overexpressing plasmids, generating long double-stranded RNA, and transfecting them into Sf9 cells. We then detail procedures for cell immunofluorescence imaging with harmine treatment and fluorescence analysis.For complete details on the use and execution of this protocol, please refer to Cui et al. (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
| first_indexed | 2024-03-11T17:24:53Z |
| format | Article |
| id | doaj.art-85e64b43dc1b4123a2ada7573dfa16c2 |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-03-11T17:24:53Z |
| publishDate | 2023-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-85e64b43dc1b4123a2ada7573dfa16c22023-10-19T04:22:51ZengElsevierSTAR Protocols2666-16672023-12-0144102646Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell linesGaofeng Cui0Zhiyan Jiang1Guohua Zhong2College of Plant Protection, South China Agricultural University, Guangzhou 510642, China; Guangdong Provincial Key Laboratory of Silviculture, Protection and Utilization, Guangdong Academy of Forestry, Guangzhou 510520, ChinaCollege of Plant Protection, South China Agricultural University, Guangzhou 510642, China; College of Food and Health, Zhejiang A&F University, Hangzhou 311300, ChinaCollege of Plant Protection, South China Agricultural University, Guangzhou 510642, China; Corresponding authorSummary: The interaction manner and biological function of Rab7 and its effector, Rab-interacting lysosomal protein (RILP), remain unclear in invertebrates. We provide a protocol for detecting the effects of Rab7 and RILP terminals on lysosome and autophagy in Spodoptera frugiperda Sf9 cells with overexpression and RNA interference. We describe steps for overexpressing plasmids, generating long double-stranded RNA, and transfecting them into Sf9 cells. We then detail procedures for cell immunofluorescence imaging with harmine treatment and fluorescence analysis.For complete details on the use and execution of this protocol, please refer to Cui et al. (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.http://www.sciencedirect.com/science/article/pii/S2666166723006135Cell BiologyMicroscopyMolecular BiologyMolecular/Chemical Probes |
| spellingShingle | Gaofeng Cui Zhiyan Jiang Guohua Zhong Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines STAR Protocols Cell Biology Microscopy Molecular Biology Molecular/Chemical Probes |
| title | Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines |
| title_full | Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines |
| title_fullStr | Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines |
| title_full_unstemmed | Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines |
| title_short | Functional assessment of lysosomal Rab7 and RILP with RNA interference and overexpression in Spodoptera frugiperda Sf9 cell lines |
| title_sort | functional assessment of lysosomal rab7 and rilp with rna interference and overexpression in spodoptera frugiperda sf9 cell lines |
| topic | Cell Biology Microscopy Molecular Biology Molecular/Chemical Probes |
| url | http://www.sciencedirect.com/science/article/pii/S2666166723006135 |
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