DamID

What is DamID?

DamID was initially developed by Bas van Steensel and Steve Henikoff. It is a method to map contact sites of proteins along the DNA. Just like chromatin immunoprecipitation, but based on an entirely different principle. DamID is based on the creation of a fusion protein consisting of Escherichia coli DNA adenine methyltransferase (Dam) and a chromatin protein or transcription factor of interest. Dam methylates adenines in the sequence GATC. Endogenous methylation of adenines is absent in most eukaryotes. Upon expression of the fusion protein in cultured cells or in an intact organism, Dam will be targeted to the native binding sites of the chromatin protein. This will then result in local methylation of adenine residues. Hence, the sequences near a binding site of the protein will be marked with a unique methylation tag, which can be detected using restriction enzymes (DpnI and DpnII) that are methylation sensitive. Protocols have been developed to combine this with next generation sequencing methods.

Drosophila DamID

We no longer do DamID in flies or fly cells, but we previously demonstrated that it can be used to map the genomic binding sites of lots of chromatin proteins and transcription factors. The vectors that we originally made for this purpose can be obtained from AddGene. Several other labs are successfully using DamID in flies, please check their papers for the latest tips and tricks.

Mammalian DamID

We only apply DamID in mammalian cells to study interactions of the genome with nuclear landmarks such as the lamina, nuclear pores, and the nucleolus. Several vectors are available from AddGene. The basic protocol can be found here, except that we and others have adapted it for Illumina sequencing (see protocol below). Other labs have also published very useful DamID-seq variants and protocols.