ORMA: a tool for identification of species-specific variations in 16S rRNA gene and oligonucleotides design

Severgnini, Marco; Cremonesi, Paola; Consolandi, Clarissa; Caredda, Giada; De Bellis, Gianluca; Castiglioni, Bianca

For each of these four options, consensus score accuracy is calculated, as the percentage of original sequences that carried the same base as the consensus in each position. (ii-iii) Design of candidate probes. We have implemented a Single Base Seeker (SBS) algorithm, for the determination of positions able to discriminate one sequence among a set of homologous ones. The discriminating position finding procedure can be summarized as follows in four basic steps: (a) Choice of a user-defined subset of sequences of the dataset (indicated as the 'positive set'). The remaining sequences are used as a group of the discriminating positions must be different from; these are addressed, in the present article, as the 'negative set'. 'Positive' and 'Negative' sets differ for the fact that every consensus in the 'positive set' group will be subjected to probe design, whereas those of the 'negative set' will not; (b) For each sequence, determination of a list of the positions of nondegenerated bases; (c) For each position on point b, calculation of a score as the sum of all the sequences carrying the same base as the considered sequence, in the same position. If the only sequence carrying the base is the tested one, the position is set as discriminating and (d) Re-calculation of the score on point c, substituting to each (eventual) degenerated base its two or three alternatives (an 'N' automatically flags the position as nondiscriminating).