Identification of LEC1 binding sites in Arabidopsis bent cotyledon stage seeds

LEAFY COTYLEDON1 (LEC1), an atypical subunit of the NF-Y CCAAT binding transcription factor, is a central regulator that controls many aspects of seed development including the maturation phase during which seeds accumulate storage macromolecules and embryos acquire the ability to withstand desiccation. To define the gene network and developmental processes controlled by LEC1, genes regulated directly by and downstream of LEC1 were identified. In this part of the study, we identified the genes bound by LEC1 in Arabidopsis bent cotyledon stage seeds. We expressed a GFP-tagged form of LEC1 under the control of its native promoter and terminator in lec1 null mutant seeds. Using an anti-GFP antibody (NeuroMab 75-131) , we performed chromatin immunoprecipitation followed by sequencing (ChIP-Seq) using chromatin isolated from bent cotyledon stage seeds to identify the genes bound by LEC1. Overall design: Plants homozygous for both the lec1-1 mutation and the LEC1:LEC1-GFP:LEC1 transgene were used to isolate chromatin from bent cotyledon-stage seeds. Anti-GFP antibodies (Neuromab 75-131) were used to immunoprecipitate LEC1-bound DNA fragments and compared to total fragmented chromatin from the same biological materials. Two biological replicates per conditions were analyzed.