Whole mount in situ hybridization detection of mRNAs using short LNA containing DNA oligonucleotide probes

Diana K. Darnell, Stacey Stanislaw, Simran Kaur, Parker B Antin

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

In situ hybridization is widely used to visualize transcribed sequences in embryos, tissues, and cells. For whole mount detection of mRNAs in embryos, hybridization with an antisense RNA probe is followed by visual or fluorescence detection of target mRNAs. A limitation of this approach is that a cDNA template of the target RNA must be obtained in order to generate the antisense RNA probe. Here we investigate the use of short (12-24 nucleotides) locked nucleic acid (LNA) containing DNA probes for whole mount in situ hybridization detection of mRNAs. Following extensive protocol optimization, we show that LNA probes can be used to localize several mRNAs of varying abundances in chicken embryos. LNA probes also detected alternatively spliced exons that are processed in a tissue specific manner. The use of LNA probes for whole mount in situ detection of mRNAs will enable in silico design and chemical synthesis and will expand the general use of in situ hybridization for studies of transcriptional regulation and alternative splicing. Published by Cold Spring Harbor Laboratory Press.

Original languageEnglish (US)
Pages (from-to)632-637
Number of pages6
JournalRNA
Volume16
Issue number3
DOIs
StatePublished - Mar 2010

Fingerprint

Oligonucleotide Probes
DNA Probes
In Situ Hybridization
Nucleic Acid Probes
Messenger RNA
RNA Probes
Antisense RNA
Embryonic Structures
Alternative Splicing
Computer Simulation
Chickens
Exons
Nucleotides
Complementary DNA
Fluorescence
locked nucleic acid
RNA

Keywords

  • Chicken embryo
  • In situ hybridization
  • Locked nucleic acids, LNA
  • mRNA detection

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Whole mount in situ hybridization detection of mRNAs using short LNA containing DNA oligonucleotide probes. / Darnell, Diana K.; Stanislaw, Stacey; Kaur, Simran; Antin, Parker B.

In: RNA, Vol. 16, No. 3, 03.2010, p. 632-637.

Research output: Contribution to journalArticle

Darnell, Diana K. ; Stanislaw, Stacey ; Kaur, Simran ; Antin, Parker B. / Whole mount in situ hybridization detection of mRNAs using short LNA containing DNA oligonucleotide probes. In: RNA. 2010 ; Vol. 16, No. 3. pp. 632-637.
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