Processes to identify variants that affect biochemical regulation are described. Generally, models are used to identify variants that affect biochemical regulation, which can be used in several downstream applications. A pathogenicity of identified variants is also determined in some instances, which can also be used in several. Various methods further develop research tools, perform diagnostics, and treat individuals based on identified variants.

The invention relates to the use of recombinant/semi-synthetic nucleosomes carrying histone and/or DNA modifications as a reference standard for quantification of covalently modified (on the histone proteins or wrapping DNA), variant, or mutant nucleosomes (collectively modified nucleosomes or nucleosome modifications) from a biological sample. The invention further relates to methods of using the assay to accurately quantify single or combinatorial nucleosome modifications as biomarkers of disease.

The invention relates to the use of recombinant/semi-synthetic nucleosomes carrying histone and/or DNA modifications as a reference standard for quantification of covalently modified (on the histone proteins or wrapping DNA), variant, or mutant nucleosomes (collectively modified nucleosomes or nucleosome modifications) from a biological sample. The invention further relates to methods of using the assay to accurately quantify single or combinatorial nucleosome modifications as biomarkers of disease.

The present invention provides methods for performing an imaging method to detect a moiety associated with labeled hydroxymethylated bases in nucleic acid sequences, such as detecting a moiety associated with glucosylated 5-hydroxmethylated cytosines.

The present invention provides methods for performing an imaging method to detect a moiety associated with labeled hydroxymethylated bases in nucleic acid sequences, such as detecting a moiety associated with glucosylated 5-hydroxmethylated cytosines.

There is described herein a method of detecting the presence of DNA from cancer cells in a subject comprising: providing a sample of cell-free DNA from a subject; subjecting the sample to library preparation to permit subsequent sequencing of the cell-free methylated DNA; adding a first amount of filler DNA to the sample, wherein at least a portion of the filler DNA is methylated, then optionally denaturing the sample; capturing cell-free methylated DNA using a binder selective for methylated polynucleotides; sequencing the captured cell-free methylated DNA; comparing the sequences of the captured cell-free methylated DNA to control cell-free methylated DNAs sequences from healthy and cancerous individuals and from individuals with distinct cancer types and subtypes; identifying the presence of DNA from cancer cells if there is a statistically significant similarity between one or more sequences of the captured cell-free methylated DNA and cell-free methylated DNAs sequences from cancerous individuals.

There is described herein a method of detecting the presence of DNA from cancer cells in a subject comprising: providing a sample of cell-free DNA from a subject; subjecting the sample to library preparation to permit subsequent sequencing of the cell-free methylated DNA; adding a first amount of filler DNA to the sample, wherein at least a portion of the filler DNA is methylated, then optionally denaturing the sample; capturing cell-free methylated DNA using a binder selective for methylated polynucleotides; sequencing the captured cell-free methylated DNA; comparing the sequences of the captured cell-free methylated DNA to control cell-free methylated DNAs sequences from healthy and cancerous individuals and from individuals with distinct cancer types and subtypes; identifying the presence of DNA from cancer cells if there is a statistically significant similarity between one or more sequences of the captured cell-free methylated DNA and cell-free methylated DNAs sequences from cancerous individuals.

The invention relates to the use of recombinant/semi-synthetic nucleosomes carrying histone and/or DNA modifications as a reference standard for quantification of covalently modified (on the histone proteins or wrapping DNA), variant, or mutant nucleosomes (collectively modified nucleosomes or nucleosome modifications) from a biological sample. The invention further relates to methods of using the assay to accurately quantify single or combinatorial nucleosome modifications as biomarkers of disease.

The invention relates to the use of recombinant/semi-synthetic nucleosomes carrying histone and/or DNA modifications as a reference standard for quantification of covalently modified (on the histone proteins or wrapping DNA), variant, or mutant nucleosomes (collectively modified nucleosomes or nucleosome modifications) from a biological sample. The invention further relates to methods of using the assay to accurately quantify single or combinatorial nucleosome modifications as biomarkers of disease.

The present invention provides for novel methods for regulating and detecting the cytosine methylation status of DNA. The invention is based upon identification of a novel and surprising catalytic activity for the family of TET proteins, namely TET1, TET2, TET3, and CXXC4. The novel activity is related to the enzymes being capable of converting the cytosine nucleotide 5-methylcytosine into 5-hydroxymethylcytosine by hydroxylation.