Whole-mount identification of gene transcripts in aphids: Protocols and evaluation of probe accessibility

Chun Che Chang, Ting Yu Huang, Chun Liang Shih, Gee Way Lin, Te Pin Chang, Hui Chiu, Wen Chang Chang

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

In situ hybridization has become a powerful tool for detecting the temporal and spatial distribution of gene transcripts in prokaryotes and eukaryotes. We report an efficient protocol for whole-mount identification of the expression of mRNAs in the parthenogenetic pea aphid Acyrthosiphon pisum, an emerging model organism with a growing accumulation of genome sequencing data. In addition to steps common for most animal in situ hybridization protocols, we describe processing methods specific to aphids, the accessibility of antisense riboprobes of different lengths in whole-mounted aphids, and signal intensity versus probe lengths. To find substrate combinations that clearly contrast single and double in situ signals in A. pisum, we tested our protocols using riboprobes constructed from two conserved germline markers, Apvasa ana Apnanos, and examined colocalized signals in the germaria and developing oocytes. Finally, we propose conditions for stringent permeabilization that may be applied to tissues deep within the aphid embryo.

Original languageEnglish
Pages (from-to)186-196
Number of pages11
JournalArchives of Insect Biochemistry and Physiology
Volume68
Issue number4
DOIs
Publication statusPublished - Aug 1 2008
Externally publishedYes

Keywords

  • Acyrthosiphon pisum
  • mRNA
  • Pea aphid
  • Whole-mount in situ hybridization

ASJC Scopus subject areas

  • Physiology
  • Biochemistry
  • Insect Science

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