Screening cDNA Libraries by Hybridization with Double-Stranded DNA Probes and Oligonucleotides

Abstract

Probably the most commonly used method to screen a cDNA library is hybridization to a labeled DNA probe. This probe may be a single-stranded oligonucleotide or a double-stranded cDNA or polymerase chain reaction (PCR) product. The DNA may be either radioactively or nonradioactively labeled. The sequence of an oligonucleotide probe may be derived from a number of sources, for example, degenerate probes may be obtained by back-translating a peptide sequence of an unknown protein, or may be a short conserved region of sequence within a cDNA from another member of a multigene family or from a cognate cDNA from another species (see Note 1). Double-stranded DNA probes may be a partial cDNA obtained by screening another library, or a PCR product, or a cDNA from another member of a gene family or from another species.