RESUMO
Re-sequencing, the identification of the specific variants in a sequence of interest compared with a known genomic sequence, is a ubiquitous task in today's biology. Universal arrays, which interrogate all possible oligonucleotides of a certain length in a target sequence, have been suggested for computationally determining a polynucleotide sequence from its oligonucleotide content. We present here new methods that use such arrays for re-sequencing. Our methods are applied to data obtained by the polymerase signaling assay, which arrays single-based primer extension reactions for either universal or partial arrays of pentanucleotides. The computational analysis uses the spectrum alignment algorithm, which is refined and enhanced here in order to overcome noise incurred by the use of such short primers. We present accurate re-sequencing results for both synthetic and amplified DNA molecules.
Assuntos
Biologia Computacional/métodos , DNA Polimerase Dirigida por DNA/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Análise de Sequência de DNA/métodos , Algoritmos , Sequência de Bases , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Variação Genética , Genótipo , Dados de Sequência Molecular , Sondas de Oligonucleotídeos/química , Alinhamento de SequênciaRESUMO
Universal arrays contain all possible oligonucleotides of a certain length, typically 6-10 bases. They can determine in a single experiment all substrings of that length that occur along a target sequence. That information, also called the spectrum of the sequence, is not sufficient to uniquely reconstruct a sequence longer than a few hundred bases. We have devised a polynomial algorithm that reconstructs the sequence, given the spectrum and an additional reference sequence, homologous to the target sequence. Such a reference is available, for example, in the identification of single-nucleotide polymorphisms. The algorithm can handle errors in the spectrum as well as substitutions, insertions, and deletions in the target sequence. We present extensive simulation results, which show that the algorithm correctly reconstructs target sequences of >2,000 nucleotides from error-prone 8-mer spectra when realistic levels of single-nucleotide polymorphisms are present.