TRAP-seq of VMH Lepr neurons

To identify transcripts enriched in ventomedial hypothalamus (VMH) Lepr neurons, we used translating affinity ribosome purification combined with RNA-seq (TRAP-seq) from Lepr-Cre mice. Lepr-Cre mice were crossed to a ROSA26 Cre-dependent EGFP-L10a line and treated with PBS or leptin. The VMH was microdissected and ribosomes were isolated using an anti-GFP-coated beads. RNA was isolated from the beads and the supernatant and converted to cDNA. Illumina libraries were prepared and sequenced on an Illumina HiSeq machine for 52 cycles.