We describe the use of in vitro DNA amplification for production of double-stranded, biotin-labeled DNA probes. Specifically, a 124 BP DNA segment of the Y chromosome-specific 3.4 KB repeat was amplified in preparations of human genomic DNA using the polymerase chain reaction (PCR) and a thermostable DNA polymerase. The PCR products were amplified further in the presence of a molar excess of biotin-11-dUTP. The resulting double-stranded DNA segments showed a high amount of incorporated biotin-11-dUTP. The probes were used in DNA-DNA hybridization experiments without further purification. When DNA sequences flanking the target region are known, probe generation by enzymatic amplification offers a rapid and efficient alternative to molecular cloning and nick translation.
- DNA probes
- In situ hybridization
- Polymerase chain reaction (PCR)
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