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      Soaking RNAi in Bombyx mori BmN4-SID1 Cells Arrests Cell Cycle Progression

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          Abstract

          RNA interference (RNAi) is an evolutionarily conserved mechanism for sequence-specific gene silencing. Previously, the BmN4-SID1 cell expressing Caenorhabditis ele gans SID-1 was established, in which soaking RNAi could induce effective gene silencing. To establish its utility, 6 cell cycle progression related cDNAs, CDK1, MYC, MYB, RNRS, CDT1, and GEMININ, were isolated from the silkworm, Bombyx mori L. (Lepidoptera: Bombycidae), and their expressions were further silenced by soaking RNAi in the BmN4-SID1 cells. The cell cycle progression analysis using flow cytometer demonstrated that the small amount of double stranded RNA was enough to arrest cell cycle progression at the specific cell phases. These data suggest that RNAi in the BmN4-SID1 cells can be used as a powerful tool for loss-of-function analysis of B. mori genes.

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          Most cited references26

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          Principles of CDK regulation.

          D Morgan (1995)
          As key regulators of the cell cycle, the cyclin-dependent kinases must be tightly regulated by extra- and intracellular signals. The activity of cyclin-dependent kinases is controlled by four highly conserved biochemical mechanisms, forming a web of regulatory pathways unmatched in its elegance and intricacy.
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            RNA interference in Lepidoptera: an overview of successful and unsuccessful studies and implications for experimental design.

            Gene silencing through RNA interference (RNAi) has revolutionized the study of gene function, particularly in non-model insects. However, in Lepidoptera (moths and butterflies) RNAi has many times proven to be difficult to achieve. Most of the negative results have been anecdotal and the positive experiments have not been collected in such a way that they are possible to analyze. In this review, we have collected detailed data from more than 150 experiments including all to date published and many unpublished experiments. Despite a large variation in the data, trends that are found are that RNAi is particularly successful in the family Saturniidae and in genes involved in immunity. On the contrary, gene expression in epidermal tissues seems to be most difficult to silence. In addition, gene silencing by feeding dsRNA requires high concentrations for success. Possible causes for the variability of success in RNAi experiments in Lepidoptera are discussed. The review also points to a need to further investigate the mechanism of RNAi in lepidopteran insects and its possible connection to the innate immune response. Our general understanding of RNAi in Lepidoptera will be further aided in the future as our public database at http://insectacentral.org/RNAi will continue to gather information on RNAi experiments. Copyright © 2010 Elsevier Ltd. All rights reserved.
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              Systemic RNAi in C. elegans requires the putative transmembrane protein SID-1.

              Double-stranded RNA-mediated gene interference (RNAi) in Caenorhabditis elegans systemically inhibits gene expression throughout the organism. To investigate how gene-specific silencing information is transmitted between cells, we constructed a strain that permits visualization of systemic RNAi. We used this strain to identify systemic RNA interference-deficient (sid) loci required to spread gene-silencing information between tissues but not to initiate or maintain an RNAi response. One of these loci, sid-1, encodes a conserved protein with predicted transmembrane domains. SID-1 is expressed in cells sensitive to RNAi, is localized to the cell periphery, and is required cell-autonomously for systemic RNAi.
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                Author and article information

                Journal
                J Insect Sci
                J. Insect Sci
                insc
                Journal of Insect Science
                University of Wisconsin Library
                1536-2442
                2013
                19 December 2013
                : 13
                : 155
                Affiliations
                [1 ]Laboratory of Silkworm Science, Kyushu University Graduate School of Bioresource and Bioenvironmental Sciences, Hakozaki 6-10-1, Fukuoka 812-8581
                [2 ]Transgenic Silkworm Research Center, National Institute of Agrobiological Sciences, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, Japan
                Author notes
                Article
                10.1673/031.013.15501
                4015410
                24773378
                ce8fea4d-148a-4f74-998d-3df5a24f1d7a
                © 2013

                This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 15 March 2012
                : 4 September 2012
                Page count
                Pages: 7
                Categories
                Article

                Entomology
                rnai,sid-1,dsrna,silkworm
                Entomology
                rnai, sid-1, dsrna, silkworm

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