High-throughput RNAi by soaking in <span class='italic'>Caenorhabtis elegans</span>

doi:10.1017/CBO9780511546402.033

30 - High-throughput RNAi by soaking in Caenorhabtis elegans

Published online by Cambridge University Press: 31 July 2009

Asako Sugimoto

Edited by

Krishnarao Appasani

Foreword by

Andrew Fire and

Marshall Nirenberg

Show author details

Asako Sugimoto: Affiliation:
Laboratory Head Laboratory for Developmental Genomics, RIKEN Center for Developmental Biology
Krishnarao Appasani: Affiliation:
GeneExpression Systems, Inc., Massachusetts
Andrew Fire: Affiliation:
Stanford University, California

Book contents

Get access

Summary

Introduction

The phenomenon of RNA-mediated interference (RNAi) was first discovered in the nematode Caenorhabditis elegans (Fire et al., 1998), in which introduction of double-stranded RNA (dsRNA) causes specific inactivation of genes with corresponding sequences. RNAi was soon recognized as an experimentally simple and technically undemanding method for gene knockdown in this organism. The emergence of this new technology, which coincided with the completion of the sequencing of the C. elegans genome (The C. elegans Sequencing Consortium, 1998), has brought about a dramatic shift in the experimental strategies adopted in the study of this organism, and greatly expanded our understanding of gene function at the whole-genome level. Our group have established an optimized “soaking method” for dsRNA delivery into worms (Maeda et al., 2001), and applied this technique for large-scale analysis of gene function. In this paper, I describe the characteristics of the “soaking method,” and its application in the study of developmental processes in C. elegans.

RNAi in C. elegans

After the finding of RNAi in C. elegans, a series of subsequent analyses demonstrated that wide range of animals, plants, and fungi has RNAi-related gene regulatory mechanisms (Cogoni and Macino, 2000). Biochemical and genetic approaches, conducted mainly in C. elegans and Drosophila, have revealed the evolutionarily conserved nature of the molecular mechanism of RNAi (see other chapters in this book for detail). Although the “core RNAi mechanism” appears to be conserved among diverse organisms, some aspects of RNAi observed in C. elegans are not universally found in other species.

Type: Chapter
Information: RNA Interference Technology
From Basic Science to Drug Development
, pp. 419 - 432

DOI: https://doi.org/10.1017/CBO9780511546402.033 [Opens in a new window]

Publisher: Cambridge University Press

Print publication year: 2005

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Austin, J. and Kimble, J. (1987). glp-1 is required in the germ line for regulation of the decision between mitosis and meiosis in C. elegans. Cell, 51, 589–599CrossRef Google Scholar PubMed

Austin, J. and Kimble, J. (1989). Transcript analysis of glp-1 and lin-12, homologous genes required for cell interactions during development of C. elegans. Cell, 58, 565–571CrossRef Google Scholar PubMed

Cogoni, C. and Macino, G. (2000). Post-transcriptional gene silencing across kingdoms. Current Opinions in Genetics and Development, 10, 638–643CrossRef Google Scholar PubMed

Dillin, A., Hsu, A. L., Arantes-Oliveira, N., Lehrer-Graiwer, J., Hsin, H., Fraser, A. G., Kamath, R. S., Ahringer, J. and Kenyon, C. (2002). Rates of behavior and aging specified by mitochondrial function during development. Science, 298, 2398–2401CrossRef Google Scholar PubMed

Elbashir, S. M., Harborth, J., Lendeckel, W., Yalcin, A., Weber, K. and Tuschl, T. (2001). Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature, 411, 494–498CrossRef Google Scholar PubMed

Fire, A., Xu, S., Montgomery, M. K., Kostas, S. A., Driver, S. E. and Mello, C. C. (1998). Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature, 391, 806–811CrossRef Google Scholar PubMed

Fraser, A. G., Kamath, R. S., Zipperlen, P., Martinez-Campos, M., Sohrmann, M. and Ahringer, J. (2000). Functional genomic analysis of C. elegans chromosome I by systematic RNA interference. Nature, 408, 325–330CrossRef Google Scholar PubMed

Gonczy, P., Echeverri, C., Oegema, K., Coulson, A., Jones, S. J., Copley, R. R., Duperon, J., Oegema, J., Brehm, M., Cassin, E., et al. (2000). Functional genomic analysis of cell division in C. elegans using RNAi of genes on chromosome III. Nature, 408, 331–336CrossRef Google Scholar

Hill, A. A., Hunter, C. P., Tsung, B. T., Tucker-Kellogg, G. and Brown, E. L. (2000). Genomic analysis of gene expression in C. elegans. Science, 290, 809–812CrossRef Google Scholar PubMed

Kamath, R. S. and Ahringer, J. (2003). Genome-wide RNAi screening in Caenorhabditis elegans. Methods, 30, 313–321CrossRef Google Scholar PubMed

Kamath, R. S., Fraser, A. G., Dong, Y., Poulin, G., Durbin, R., Gotta, M., Kanapin, A., Bot, N., Moreno, S., Sohrmann, M., et al. (2003). Systematic functional analysis of the Caenorhabditis elegans genome using RNAi. Nature, 421, 231–237CrossRef Google Scholar PubMed

Kamath, R. S., Martinez-Campos, M., Zipperlen, P., Fraser, A. G., and Ahringer, J. (2001). Effectiveness of specific RNA-mediated interference through ingested double-stranded RNA in Caenorhabditis elegans. Genome Biology, 2, RESEARCH0002Google Scholar PubMed

Kelly, W. G., Schaner, C. E., Dernburg, A. F., Lee, M. H., Kim, S. K., Villeneuve, A. M. and Reinke, V. (2002). X-chromosome silencing in the germline of C. elegans. Development, 129, 479–492Google Scholar PubMed

Kim, S. K., Lund, J., Kiraly, M., Duke, K., Jiang, M., Stuart, J. M., Eizinger, A., Wylie, B. N. and Davidson, G. S. (2001). A gene expression map for Caenorhabditis elegans. Science, 293, 2087–2092CrossRef Google Scholar PubMed

Kuroyanagi, H., Kimura, T., Wada, K., Hisamoto, N., Matsumoto, K. and Hagiwara, M. (2000). SPK-1, a C. elegans SR protein kinase homologue, is essential for embryogenesis and required for germline development. Mechanisms of Development, 99, 51–64CrossRef Google Scholar

Lee, S. S., Lee, R. Y., Fraser, A. G., Kamath, R. S., Ahringer, J. and Ruvkun, G. (2003). A systematic RNAi screen identifies a critical role for mitochondria in C. elegans longevity. Nature Genetics, 33, 40–48CrossRef Google Scholar PubMed

Li, S., Armstrong, C. M., Bertin, N., Ge, H., Milstein, S., Boxem, M., Vidalain, P. O., Han, J. D., Chesneau, A., Hao, T., Goldberg, D. S., Li, N., Martinez, M., Rual, J. F., Lamesch, P., Xu, L., Tewari, M., Wong, S. L., Zhang, L. V., Berriz, G. F., Jacotot, L., Vaglio, P., Reboul, J., Hirozane-Kishikawa, T., Li, Q., Gabel, H. W., Elewa, A., Baumgartner, B., Rose, D. J., Yu, H., Bosak, S., Sequerra, R., Fraser, A., Mango, S. E., Saxton, W. M., Strome, S., Heuvel, S., Piano, F., Vandenhaute, J., Sardet, C., Gerstein, M., Doucette-Stamm, L., Gunsalus, K. C., Harper, J. W., Cusick, M. E., Roth, F. P., Hill, D. E. and Vidal, M. (2004). A Map of the Interactome Network of the Metazoan C. elegans. Science, 303, 540–543CrossRef Google Scholar PubMed

Maeda, I., Kohara, Y., Yamamoto, M. and Sugimoto, A. (2001). Large-scale analysis of gene function in Caenorhabditis elegans by high-throughput RNAi. Current Biology, 11, 171–176CrossRef Google Scholar PubMed

Montgomery, M. K., Xu, S. and Fire, A. (1998). RNA as a target of double-stranded RNA-mediated genetic interference in Caenorhabditis elegans. Proceedings of the National Academy of Sciences USA, 95, 15502–15507CrossRef Google Scholar PubMed

Paddison, P. J., Caudy, A. A., Bernstein, E., Hannon, G. J. and Conklin, D. S. (2002). Short hairpin RNAs (shRNAs) induce sequence-specific silencing in mammalian cells. Genes & Development, 16, 948–958CrossRef Google Scholar PubMed

Piano, F., Schetter, A. J., Mangone, M., Stein, L. and Kemphues, K. J. (2000). RNAi analysis of genes expressed in the ovary of Caenorhabditis elegans. Current Biology, 10, 1619–1622CrossRef Google Scholar PubMed

Piano, F., Schetter, A. J., Morton, D. G., Gunsalus, K. C., Reinke, V., Kim, S. K. and Kemphues, K. J. (2002). Gene clustering based on RNAi phenotypes of ovary-enriched genes in C. elegans. Current Biology, 12, 1959–1964CrossRef Google Scholar PubMed

Priess, J. R., Schnabel, H. and Schnabel, R. (1987). The glp-1 locus and cellular interactions in early C. elegans embryos. Cell, 51, 601–611CrossRef Google Scholar PubMed

Reboul, J., Vaglio, P., Rual, J. F., Lamesch, P., Martinez, M., Armstrong, C. M., Li, S., Jacotot, L., Bertin, N., Janky, R., et al. (2003). C. elegans ORFeome version 1.1: Experimental verification of the genome annotation and resource for proteome-scale protein expression. Nature Genetics, 34, 35–41CrossRef Google Scholar PubMed

Reboul, J., Vaglio, P., Tzellas, N., Thierry-Mieg, N., Moore, T., Jackson, C., Shin-i, T., Kohara, Y., Thierry-Mieg, D., Thierry-Mieg, J., et al. (2001). Open-reading-frame sequence tags (OSTs) support the existence of at least 17300 genes in C. elegans. Nature Genetics, 27, 332–336CrossRef Google Scholar PubMed

Reinke, V., Smith, H. E., Nance, J., Wang, J., Doren, C., Begley, R., Jones, S. J., Davis, E. B., Scherer, S., Ward, S. and Kim, S. K. (2000). A global profile of germline gene expression in C. elegans. Molecular Cell, 6, 605–616CrossRef Google Scholar PubMed

Sijen, T., Fleenor, J., Simmer, F., Thijssen, K. L., Parrish, S., Timmons, L., Plasterk, R. H. and Fire, A. (2001). On the role of RNA amplification in dsRNA-triggered gene silencing. Cell, 107, 465–476CrossRef Google Scholar PubMed

Simmer, F., Moorman, C., Linden, A. M., Kuijk, E., Berghe, P. V., Kamath, R., Fraser, A. G., Ahringer, J. and Plasterk, R. H. (2003). Genome-wide RNAi of C. elegans using the hypersensitive rrf-3 strain reveals novel gene functions. Public Library of Science Biology, 1, E12Google Scholar PubMed

Simmer, F., Tijsterman, M., Parrish, S., Koushika, S. P., Nonet, M. L., Fire, A., Ahringer, J. and Plasterk, R. H. (2002). Loss of the putative RNA-directed RNA polymerase RRF-3 makes C. elegans hypersensitive to RNAi. Current Biology, 12, 1317–1319CrossRef Google Scholar PubMed

Stein, L., Sternberg, P., Durbin, R., Thierry-Mieg, J. and Spieth, J. (2001). Worm Base: network access to the genome and biology of Caenorhabditis elegans. Nucleic Acids Research, 29, 82–86CrossRef Google Scholar

Tabara, H., Grishok, A. and Mello, C. C. (1998). RNAi in C. elegans: Soaking in the genome sequence. Science, 282, 430–431CrossRef Google Scholar

Tavernarakis, N., Wang, S. L., Dorovkov, M., Ryazanov, A. and Driscoll, M. (2000). Heritable and inducible genetic interference by double-stranded RNA encoded by transgenes. Nature Genetics, 24, 180–183CrossRef Google Scholar PubMed

The C. elegans Sequencing Consortium. (1998). Genome sequence of the nematode C. elegans: A platform for investigating biology. Science, 282, 2012–2018CrossRef

Timmons, L. and Fire, A. (1998). Specific interference by ingested dsRNA. Nature, 395, 854CrossRef Google Scholar PubMed

Winston, W. M., Molodowitch, C. and Hunter, C. P. (2002). Systemic RNAi in C. elegans requires the putative transmembrane protein SID-1. Science, 295, 2456–2459CrossRef Google Scholar

Book contents

30 - High-throughput RNAi by soaking in Caenorhabtis elegans

Summary

Access options

References

Save book to Kindle

Save book to Dropbox

Save book to Google Drive