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Targeted gene walking polymerase chain reaction
J D Parker1, P S Rabinovitch, G C Burmer
1Department of Pathology, University of Washington, Seattle 98195.
Nucleic Acids Research
|June 11, 1991
Summary
Targeted gene walking, a modified polymerase chain reaction (PCR) technique, amplifies unknown DNA sequences next to known DNA. This method expedites DNA sequence isolation, replacing traditional cloning and screening steps.
Area of Science:
- Molecular Biology
- Genetics
- Biotechnology
Background:
- Traditional methods for isolating unknown DNA sequences adjacent to known sequences often involve laborious cloning and screening processes.
- These conventional techniques can be time-consuming and inefficient, hindering rapid genetic analysis and discovery.
Purpose of the Study:
- To introduce and describe a modified polymerase chain reaction (PCR) technique named 'targeted gene walking'.
- To demonstrate the utility of targeted gene walking as a rapid, single-step alternative to conventional cloning and screening for DNA sequence isolation.
Main Methods:
- Targeted gene walking utilizes a combination of one sequence-specific PCR primer and one non-specific 'walking' primer.
- This approach enables the amplification of unknown DNA sequences flanking a known DNA region.
Main Results:
- The targeted gene walking technique significantly expedites the isolation of DNA sequences upstream or downstream from a known sequence.
- It offers a streamlined, single-step PCR protocol, reducing the time and effort compared to traditional methods.
Conclusions:
- Targeted gene walking is an effective method for rapidly isolating unknown contiguous DNA sequences.
- This technique has broad applications in gene discovery, polymorphism detection, and regulatory region identification.