title: Real-time PCR mapping of DNaseI-hypersensitive sites using a novel ligation-mediated amplification technique.
abstract: Mapping sites within the genome that are hypersensitive to digestion with DNaseI is an important method for identifying DNA elements that regulate transcription. The standard approach to locating these DNaseI-hypersensitive sites (DHSs) has been to use Southern blotting techniques, although we, and others, have recently published alternative methods using a range of technologies including high-throughput sequencing and genomic array tiling paths. In this article, we describe a novel protocol to use real-time PCR to map DHS. Advantages of the technique reported here include the small cell numbers required for each analysis, rapid, relatively low-cost experiments with minimal need for specialist equipment. Presented examples include comparative DHS mapping of known TAL1/SCL regulatory elements between human embryonic stem cells and K562 cells.
authors: Department of Haematology, Cambridge Institute for Medical Research, Addenbrooke's Hospital, University of Cambridge, Cambridge CB2 2XY, UK. gf246@cam.ac.uk
authors: George A
authors: GA
authors: Follows
authors: Mary E
authors: ME
authors: Janes
authors: Ludovic
authors: L
authors: Vallier
authors: Anthony R
authors: AR
authors: Green
authors: Berthold
authors: B
authors: Gottgens
mesh_terms: D015139:Blotting, Southern; D002478:Cells, Cultured; D002874:Chromosome Mapping / Q000379:methods*; D004247:DNA / Q000032:analysis; D003850:Deoxyribonuclease I; D053595:Embryonic Stem Cells / Q000737:chemistry; D015698:Genomic Library; D006801:Humans; D020014:K562 Cells; D015459:Leukemia-Lymphoma, Adult T-Cell / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D012045:Regulatory Sequences, Nucleic Acid
chemical_list: D004247:DNA; D003850:Deoxyribonuclease I