Terminal Restriction Fragment Length Polymorphism

T-RFLP (Terminal Restriction fragment Length Polymorphism) is one of the most common DNA fingerprinting techniques in forensic applications. T-RFLP is a microbial profiling method and is an amplification method for amplifying the 16S rRNA genes. TRFLP is an amplification method using fluorescent-labeled end primer.

Terminal Restriction Fragment Length Polymorphism

Amplicons are digested using restriction endonucleases. Fragments (different sizes) are separated by electrophoresis. After electrophoresis terminal fragments can be visualized. The amplified fragments originate from different organisms and have different sequence variations. Terminal restriction sites for different species are unique and aids in differentiating microbial profiles.

Terminal Restriction Fragment Length Polymorphism

Terminal Restriction Fragment Length Polymorphism patterns are used to characterize the microbial communities from sites contaminated with a pollutant and are compared with the microbial community profile from uncontaminated areas of the site in the above picture. The profile of the microbial community contains a number of peaks in clean groundwater indicative of a complex community. The presence of benzene in the profiles (b and c) is indicated by the presence of fewer peaks which indicates the community is dominated by fewer micro-organisms as a result of benzene contamination. Microbial communities can be monitored using this technique and enable greater reproducibility.

The disadvantage of this technique is that sample is destructively sampled so that DNA cannot be used again unlike in Denaturing Gradient Gel Electrophoresis (DGGE) which allows for either direct cloning of bands or the direct sequencing of bands excised from the gel.

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