Comparative analysis of microsatellite detecting software: a significant variation in results and influence of parameters

Microsatellites are a unique type of repeat patterns found in genome sequences of all known organisms including bacteria and viruses. These repeats play an important role in genome evolution, are associated with various diseases, have been used as molecular markers in DNA Fingerprinting, Population Genetics etc. Various bioinformatics tools have been developed for extraction of microsatellites from DNA sequences. However, not all tools can identify microsatellites with similar sensitivities and hence studies on microsatellites can suffer from significant biases in results and interpretations depending on the type of tool used. In order to get a clear idea on inherent limitations and biases with regard to extraction of microsatellites especially under the influence of varying threshold values of program parameters we carried out a comparative analysis of performance of some of the widely used tools using some test DNA sequences. We extracted imperfect microsatellites from three different sequences (E. coli bacterial genome, C. elegans Chromosome I and Drosophila Chromosome X) using the commonly used microsatellite extraction tools TRF, Sputnik, SciRoKoCo and IMEx with varying parameters and analyzed the results. We observed a significant variation in the number of microsatellites extracted by these tools even when used with default / suggested parameters. Relaxation of parameter values lead to an increase in the number of repeats detected but still the differences among the results persist. In TRF, Sputnik and SciRoKoCo it was observed that the number of mismatches increases with the increase in the tract length of the repeat indicating the level of imperfection is not uniform throughout the repeats. The four tools investigated in this study differ in their algorithms, in the parameters they use and hence in the number of microsatellites detected. The score based programs identify more number of divergent penta and hexa nucleotide repeats than IMEx. We therefore suggest that it is prudent to alter parameters appropriately to detect as many microsatellites as possible as a means not to miss any genuine repeat tracts or to use more than one tool as a means to get a good consensus. We also made a detailed survey of the available features of all microsatellite extraction tools. Apart from differences in their algorithm, efficiency and parameters, the tools also differ largely in terms of the features and flexibility.