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

Fingerprint

Aphids
Acyrthosiphon pisum
probes (equipment)
Aphidoidea
Genes
in situ hybridization
Spatial distribution
In Situ Hybridization
Animals
genes
Anas
Tissue
prokaryotic cells
processing technology
Messenger RNA
eukaryotic cells
germ cells
Peas
embryo (animal)
oocytes

Keywords

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

ASJC Scopus subject areas

  • Physiology
  • Biochemistry
  • Insect Science

Cite this

Whole-mount identification of gene transcripts in aphids : Protocols and evaluation of probe accessibility. / Chang, Chun Che; Huang, Ting Yu; Shih, Chun Liang; Lin, Gee Way; Chang, Te Pin; Chiu, Hui; Chang, Wen Chang.

In: Archives of Insect Biochemistry and Physiology, Vol. 68, No. 4, 01.08.2008, p. 186-196.

Research output: Contribution to journalArticle

Chang, Chun Che ; Huang, Ting Yu ; Shih, Chun Liang ; Lin, Gee Way ; Chang, Te Pin ; Chiu, Hui ; Chang, Wen Chang. / Whole-mount identification of gene transcripts in aphids : Protocols and evaluation of probe accessibility. In: Archives of Insect Biochemistry and Physiology. 2008 ; Vol. 68, No. 4. pp. 186-196.
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