Subtractive hybridization strategy using paramagnetic oligo(dT) beads and PCR

Melinda Mészáros, David Morton

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Subtractive hybridization has been widely used for the identification of differentially expressed genes. Here we describe a simple, sensitive strategy of subtractive hybridization that involves binding the driver poly(A)+ RNA pool to paramagnetic Dynabeads® Oligo (dT)25. After hybridization with target cDNA, the molecules common to both pools are removed. The subtracted cDNA is then amplified with PCR and used for library screening. Using this method, we have identified four cDNA clones that represent developmentally regulated transcripts in the central nervous system of the tobacco hornworm Manduca sexta. All four transcripts are of low abundance, comprising only 0.001%-0.5% of the poly(A)+ RNA pool.

Original languageEnglish (US)
Pages (from-to)413-419
Number of pages7
JournalBioTechniques
Volume20
Issue number3
StatePublished - Mar 1996
Externally publishedYes

Fingerprint

Manduca
Complementary DNA
Polymerase Chain Reaction
Messenger RNA
Tobacco
Neurology
Screening
Central Nervous System
Clone Cells
Genes
Molecules
Subtractive Hybridization Techniques
oligo (dT)

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Subtractive hybridization strategy using paramagnetic oligo(dT) beads and PCR. / Mészáros, Melinda; Morton, David.

In: BioTechniques, Vol. 20, No. 3, 03.1996, p. 413-419.

Research output: Contribution to journalArticle

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