Using RNAi to investigate orthologous homeotic gene function during development of distantly related insects

Brown, Susan J.; Mahaffey, James W.; Lorenzen, Marcé D.; Denell, Robin E.

doi:10.1046/j.1525-142x.1999.99013.x

Cited by 142 publications

(117 citation statements)

References 26 publications

(29 reference statements)

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“…In the absence of genetic tools that allow us to establish gene functions in Megaselia, we adopted the RNAi technique to assess the function of Ma-bcd and Ma-hb. Injection of dsRNA into insect embryos has already been demonstrated to cause phenocopies of zygotically expressed segmentation genes (39)(40)(41). However, genes that are expressed and required earlier than the genes of the zygotic segmentation cascade, such as maternally derived bicoid or maternally and zygotically expressed hunchback, have not yet been examined by this technique.…”

Section: Rnai Induced Phenocopies Of Bicoid and Hunchback Mutations Inmentioning

confidence: 99%

Function of bicoid and hunchback homologs in the basal cyclorrhaphan fly Megaselia (Phoridae)

Stauber

Taubert

Schmidt‐Ott

2000

Proc. Natl. Acad. Sci. U.S.A.

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The Drosophila gene bicoid functions at the beginning of a gene cascade that specifies anterior structures in the embryo. Its transcripts are localized at the anterior pole of the oocyte, giving rise to a Bicoid protein gradient, which regulates the spatially restricted expression of target genes along the anterior-posterior axis of the embryo in a concentration-dependent manner. The morphogen function of Bicoid requires the coactivity of the zinc finger transcription factor Hunchback, which is expressed in a Bicoid-dependent fashion in the anterior half of the embryo. Whereas hunchback is conserved throughout insects, bicoid homologs are known only from cyclorrhaphan flies. Thus far, identification of hunchback and bicoid homologs rests only on sequence comparison. In this study, we used double-stranded RNA interference (RNAi) to address the function of bicoid and hunchback homologs in embryos of the lower cyclorrhaphan fly Megaselia abdita (Phoridae). Megaselia-hunchback RNAi causes hunchback-like phenotypes as observed in Drosophila, but Megaselia-bicoid RNAi causes phenotypes different from corresponding RNAi experiments in Drosophila and bicoid mutant embryos. Megaselia-bicoid is required not only for the head and thorax but also for the development of four abdominal segments. This difference between Megaselia and Drosophila suggests that the range of functional bicoid activity has been reduced in higher flies.

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Section: Rnai Induced Phenocopies Of Bicoid and Hunchback Mutations Inmentioning

confidence: 99%

Function of bicoid and hunchback homologs in the basal cyclorrhaphan fly Megaselia (Phoridae)

Stauber

Taubert

Schmidt‐Ott

2000

Proc. Natl. Acad. Sci. U.S.A.

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“…Injecting dsRNA into larvae allows functional analysis at a variety of beetle life stages, including the larval, pupal, and adult stages 4,5,10 . We have previously reported our overall larval RNAi protocol including molecular biology procedures 11 .…”

Section: Introductionmentioning

confidence: 99%

Larval RNA Interference in the Red Flour Beetle, <em>Tribolium castaneum</em>

Linz

Clark-Hachtel

Borràs-Castells

et al. 2014

JoVE

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The red flour beetle, Tribolium castaneum, offers a repertoire of experimental tools for genetic and developmental studies, including a fully annotated genome sequence, transposon-based transgenesis, and effective RNA interference (RNAi). Among these advantages, RNAi-based gene knockdown techniques are at the core of Tribolium research. T. castaneum show a robust systemic RNAi response, making it possible to perform RNAi at any life stage by simply injecting double-stranded RNA (dsRNA) into the beetle's body cavity.In this report, we provide an overview of our larval RNAi technique in T. castaneum. The protocol includes (i) isolation of the proper stage of T. castaneum larvae for injection, (ii) preparation for the injection setting, and (iii) dsRNA injection. Larval RNAi is a simple, but powerful technique that provides us with quick access to loss-of-function phenotypes, including multiple gene knockdown phenotypes as well as a series of hypomorphic phenotypes. Since virtually all T. castaneum tissues are susceptible to extracellular dsRNA, the larval RNAi technique allows researchers to study a wide variety of tissues in diverse contexts, including the genetic basis of organismal responses to the outside environment. In addition, the simplicity of this technique stimulates more student involvement in research, making T. castaneum an ideal genetic system for use in a classroom setting.

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“…From its initial use in 1999 to phenocopy null mutations of the T. castaneum Deformed gene (Brown et al, 1999), to its wide use in functional genomic screens, RNAi has become an extremely valuable tool both to elucidate gene function and to identify potential pest control targets. Here we will discuss key Tribolium-based RNAi studies with significance to pest control.…”

Section: Red Flour Beetlementioning

confidence: 99%