JBP1-Seq
JBP1-Seq
J-binding protein sequencing (JBP1-seq) method relies on the strong affinity the J-binding protein 1 (JBP1) has for 5-ghmC. A recombinant JBP1 protein with an additional His tag and Avi tag was biotinylated in vivo. After protein purification, the biotinylated JBP1 was conjugated to streptavidin-coated magnetic beads.
The four main steps of the JBP1-seq workflow are: Tagmentation of the genomic DNA to simultaneously fragment DNA and add adapter sequences. The 5hmC sites are glucosylated by T4-βGT and the JBP1-magnetic beads used to enrich for fragments containing β-glu-5hmC. The enriched library was amplified to add on the P5 and P7 adaptors and barcodes for sequencing.