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Small Molecule Injection into Single-Cell C. elegans Embryos via Carbon-Reinforced Nanopipettes

Cornell Affiliated Author(s)

Author

L.D. Brennan
T. Roland
D.G. Morton
S.M. Fellman
S.Y. Chung
M. Soltani
J.W. Kevek
P.M. McEuen
K.J. Kemphues
M.D. Wang

Abstract

The introduction of chemical inhibitors into living cells at specific times in development is a useful method for investigating the roles of specific proteins or cytoskeletal components in developmental processes. Some embryos, such as those of Caenorhabditis elegans, however, possess a tough eggshell that makes introducing drugs and other molecules into embryonic cells challenging. We have developed a procedure using carbon-reinforced nanopipettes (CRNPs) to deliver molecules into C. elegans embryos with high temporal control. The use of CRNPs allows for cellular manipulation to occur just subsequent to meiosis II with minimal damage to the embryo. We have used our technique to replicate classical experiments using latrunculin A to inhibit microfilaments and assess its effects on early polarity establishment. Our injections of latrunculin A confirm the necessity of microfilaments in establishing anterior-posterior polarity at this early stage, even when microtubules remain intact. Further, we find that latrunculin A treatment does not prevent association of PAR-2 or PAR-6 with the cell cortex. Our experiments demonstrate the application of carbon-reinforced nanopipettes to the study of one temporally-confined developmental event. The use of CRNPs to introduce molecules into the embryo should be applicable to investigations at later developmental stages as well as other cells with tough outer coverings. © 2013 Brennan et al.

Date Published

Journal

PLoS ONE

Volume

8

Issue

9

URL

https://www.scopus.com/inward/record.uri?eid=2-s2.0-84884608403&doi=10.1371%2fjournal.pone.0075712&partnerID=40&md5=ca975fbe94748a8331785e5e54a1e16f

DOI

10.1371/journal.pone.0075712

Research Area

Group (Lab)

Michelle Wang Group
Paul McEuen Group

Funding Source

GM059849
GM79112
MCB-0820293
P40 OD010440
R01GM059849

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