Profiling method

Abstract

A polynucleotide comprising an indexing barcode incorporated at a first end of the polynucleotide only and a label bound site-specifically to a nucleotide. Preferably the nucleotide is between 100 – 500 nucleotides and the label is not fluorescent The label comprises a tag from a cofactor analogue, a high affinity binding protein or antibody or a DNA binding protein that has been crosslinked to a nucleotide residue Also disclosed is an amplified sequencing library comprising a first indexing barcode, a first subset of polynucleotides comprising a site-specifically bound label and a second subset of unlabelled polynucleotides. Further disclosed is a method of preparing a polynucleotide sample for sequencing, where the polynucleotide is suitable for determining the status of nucleotide residues and the presence of a genetic mutation The label is a methyl-CpG binding domain protein, a high affinity antibody specific for, 5-methylcytosine, 5-hydroxymethy[cytosine, N4-methylcytosine or N6-methylcytosine. Further disclosed is an in vitro method of diagnosing a disease in a subject, based on the profile obtained by the method and a kit for determining the status of nucleotide residues and the presence of a genetic mutation in a polynucleotide sample.