T-RFLP

T-RFLP Terminal restriction fragment length polymorphism... 

Covert and Moran (2002) Marine Whole, HMW, and LMW DOM Bacterial respiration 16S rDNA clone libraries and T-RFLP No 6 weeks LMW DOM 4-1 Ox more bioavailable HMW DOM community dominated by a- and p-Proteobacteria LMW DOM community dominated by y- and 8-Proteobacteria... 

Marsh, T. L. 1999. Terminal restriction fragment length polymorphism (T-RFLP) An emerging method for characterizing diversity among homologous populations of amplification products. Current Opinion in Microbiology 2 323-327. 

Osborn, A. M., Moore, E. R. B., and Timmis, K. N. (2000). An evaluation of terminal-restriction fragment length polymorphism (T-RFLP) analysis for the study of microbial community structure and dynamics. Environ. Microbiol. 2,39-50. 

Dickie, I. A., Xu, B. Koide, R. T. (2002). Vertical niche differentiation of ectomycorrhizal hyphae in soil as shown by T-RFLP analysis. New Phytologist, 156, 527-35. 

Horz, H. P., Rotthauwe, J. H., Lukow, T., and Liesack, W. (2000). Identification of major subgroups of ammonia-oxidizing bacteria in environmental samples by T-RFLP analysis of amoA PCR products. J. Microbiol. Meth. 39, 197—204. 

Detail microbial study is crucial to correlate hydrogen production to the microbial group involved. For this purpose, a nucleic acid based technique - terminal restriction fragment length polymorphism (T-RFLP) - was used to identify the... 

Terminal Restriction Fragment Length Polymorphism (T-RFLP)... 

T-RFLP has been used to study microbial diversity in contaminated sediments and to monitor changes in community composition following exposure to pollutants. In a study by Covert and Moran [77], the structure of bacterial communities growing at high-molecular weight (HMW) and at low-molecular weight (LMW) dissolved organic carbon was compared by T-RFLP. 

Fig. 7 Principle of the T-RFLP technique. Following extraction of nucleic acids, PGR amplification is performed using one fluorescently labeled primer. The amplified products are then subjected to restriction digestion, which leads to the generation of fluorescently labeled fragments of different lengths. The fragments are separated on a sequencing gel...

The results showed a dominance of gamma and epsilon Proteobacteria in the case of LMW but a dominance of alpha, beta, and gamma Proteobacteria in addition to members of the Cytophaga/Flavobacterium/Bacteriodes group in the case of HMW. Konstantinidis et al. [78] used T-RFLP in order to estimate the bacterial diversity in sediments contaminated with heavy metals. Their analysis revealed a conserved community structure over a depth profile of 15 cm. 

T-RFLP could also be used to follow shifts in bacterial community structure. Turpeinen et al. [79], used this technique to study the impact of contamination by arsenic, chromium and copper on microbial communities. The T-RFLP analysis showed that contamination with metals induces permanent changes in the microbial community structure. Rousseaux et al. [80] also re-... 

T-RFLP has been used to identify bacteria hving in consortia. An example of this is the identification of an aerobic microbial consortium that reductively dechlorinates trichloroethene completely to ethane [82]. T-RFLP patterns suggested that the consortium was dominated by populations belonging to three phylogenetic groups Dehalococcoid.es, Desulfovibrio and members of the Clostridiaceae. This composition was further confirmed by cloning. 

T-RFLP has several limitations due to the preferential extraction of DNA and PCR amplification biases. These problems have been discussed in detail in [84,85]. Another problem is the incomplete and non-specific restriction digestion. However, this can be solved by including an amplified product from... 

T-RFLP High-throughput, sensitive, and automatable process Pseudo-quantitative method Expensive equipment... 

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