Technology
A molecular genetic technique that isolates and sequences chromosomal regions that are rich in open chromatin. First, nuclei are harvested from a cellular sample. Then a hyperactive Tn5 transposase is added to the nuclei where it excises non-nucleosomal DNA strands and ligates co-administered high-throughput sequencing adapters (tagmentation). The tagged DNA fragments are isolated, amplified by PCR and sequenced. The number of reads for specific region of DNA correlate with increased chromatin accessibility and this method can identify regions of transcription factor and nucleosome binding.
Diagnostic Methods
Technology
A DNA sequencing technique that can differentiate cytosine from 5-methylcytosine in a DNA sample. First, a denatured DNA sample is treated with bisulfite which converts non-methylated cytosine to uracil. Next, the sample is amplified using a PCR method that does not discriminate between non-methylated and methylated sequences. The amplified DNA is subjected to nucleotide sequencing. The resulting sequence is compared to an identical control sample of DNA that was not treated with bisulfite. Unmethylated cytosines will be displayed as cytosines in the control sample and as thymines in the bisulfite-treated sample.
Diagnostic Methods
Technology
A molecular genetic technique that combines chromatin immunoprecipitation (ChIP) with massively parallel DNA sequencing to map the binding sites of DNA-associated proteins in a sample of cells. First, crosslinked protein-DNA complexes are isolated using ChIP. Next, the crosslinks are broken, the proteins are removed and the purified DNA is modified with adaptor oligonucleotides to facilitate massively parallel DNA sequencing. Following sequencing, the DNA sequences that are obtained can be mapped to their genomic locations.
Diagnostic Methods
Technology
A technique that utilizes immunoprecipitation to find protein-DNA interactions in a biological sample. First, the sample is treated causing the formation of protein-chromatin crosslinks. Then, the sample is sheared to separate individual protein-DNA complexes. Finally, an antibody directed against a protein of interest is applied and the protein-DNA complexes are immunoprecipitated.
Diagnostic Methods
Technology
Comparative genomic hybridization (CGH) is a technique that allows the detection of losses and gains in DNA copy number across the entire genome without prior knowledge of specific chromosomal abnormalities. Comparative genomic hybridization utilizes the hybridization of differentially labeled tumor and reference DNA to generate a map of DNA copy number changes in tumor genomes. Comparative genomic hybridization is an ideal tool for analyzing chromosomal imbalances in archived tumor material and for examining possible correlations between these findings and tumor phenotypes. (from Ried et al. J Mol Med 1997 75:801-14)
Diagnostic Methods
Technology
A molecular biology technique used to detect the interaction of a DNA binding protein with its cognate binding sequence. Labeled DNAs were reacted with crude cell extracts and the complexes are run through a non-denaturing polyacrylamide gel. The migration of the labeled DNA through the gel will be slower by being bound. This shift in electrophoretic mobility indicates functional binding between the protein and the DNA.
Diagnostic Methods
Technology
Gel electrophoresis is a method of separating large molecules (such as DNA fragments or proteins) from a mixture of similar molecules by electrophoretic filtration through a gelatinous matrix. Subject to an electric field, each molecule in the sample mixture migrates through the matrix at a different rate, depending mainly on electrical charge and size, resulting in spatial separation within the matrix of each molecular species in the sample mixture. Agarose and acrylamide in a cylindrical tube or slab gel are commonly used media for electrophoresis of proteins and nucleic acids.
Diagnostic Methods
Technology
Use of a DNA or RNA probe to detect the presence of complementary sequences in chromosomes, cells, or tissues.
Diagnostic Methods
Technology
A type of nucleic acid amplification test in which sample DNA is exposed to a single temperature in the presence of multiple sets of primers and a polymerase with replication and high strand displacement activity. This test negates the need for thermal cyclers and may be a low cost alternative to PCR.
Diagnostic Methods
Technology
An analysis done to detect monoallelic loss of genes or DNA regions in a diploid cell.
Diagnostic Methods
Technology
An analysis of DNA or protein to detect the presence and/or location of methyl groups.
Diagnostic Methods
Technology
A type of high-throughput, multiplex PCR method used to determine abnormal DNA copy number changes of up to 50 genomic DNA or RNA sequences with a single pair of PCR primers. It is frequently used in the detection of genetic aberrations in tumors, and can distinguish sequences differing in only a single nucleotide.
Diagnostic Methods
