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Technicalities and Glitches of Terminal Restriction Fragment Length Polymorphism (T-RFLP)

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Abstract

Terminal restriction fragment length polymorphism (T-RFLP) is a rapid, robust, inexpensive and simple tool for microbial community profiling. Methods used for DNA extraction, PCR amplification and digestion of amplified products have a considerable impact on the results of T-RFLP. Pitfalls of the method skew the similarity analysis and compromise its high throughput ability. Despite a high throughput method of data generation, data analysis is still in its infancy and needs more attention. Current article highlights the limitations of the methods used for data generation and analysis. It also provides an overview of the recent methodological developments in T-RFLP which will assist the readers in obtaining real and authentic profiles of the microbial communities under consideration while eluding the inherent biases and technical difficulties.

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Acknowledgments

The authors are thankful to the Department of Biotechnology (DBT), Government of India for supporting this work by grant BT/PR10054/NDB/52/94/2007. The review benefited from valuable inputs and comments from Prof. Andrew Ogram, Soil and Water Science Department, University of Florida, USA, and Somak Chowdhury and Shreyas Kumbhare from National Centre for Cell Science, Pune, India.

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  1. Microbial Culture Collection, National Centre for Cell Science, NCCS Complex, Ganeshkhind, Pune, 411007, Maharashtra, India

    Om Prakash, Prashant K. Pandey, Girish J. Kulkarni, Kiran N. Mahale & Yogesh S. Shouche

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  1. Om Prakash
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  2. Prashant K. Pandey
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Correspondence to Om Prakash.

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Prakash, O., Pandey, P.K., Kulkarni, G.J. et al. Technicalities and Glitches of Terminal Restriction Fragment Length Polymorphism (T-RFLP). Indian J Microbiol 54, 255–261 (2014). https://doi.org/10.1007/s12088-014-0461-0

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