Genome-wide occupancy profiling of Ronin (Thap11) in retinal development. Mus musculus

Conditional knockout of the transcription factor Ronin (Thap11) in retinal progenitor cells (RPCs) results in a profound failure cell proliferation resulting in a hypoplastic adult retina that also suffers from photoreceptor degeneration. The goal of this study was to determine the genes that are transcriptionally regulated by Ronin during retinogenesis. P0 wild type retinae (CD-1 background) were pooled (>10 each) in ice-cold 1X PBS and immediately processed for chromatin extraction, fragmentation and immunoprecipitation using custom antibodies against Ronin G4275 (Dejosez et al., 2010), G4275 preimmune serum or normal rabbit IgG (Santa Cruz, sc-2027). The immunoprecipitated DNA fragments were then sequenced using the Ion Torrent PGM system. Overall design: Examination of Ronin binding sites throughout the postnatal day 0 (P0) retinal genome.

转录因子Ronin（Thap11）在视网膜祖细胞（retinal progenitor cells, RPCs）中的条件性敲除，会导致细胞增殖严重受阻，最终形成发育不全的成年视网膜，同时还会伴随光感受器变性。本研究旨在鉴定视网膜发生过程中受Ronin转录调控的基因。取背景为CD-1品系的出生后第0天（P0）野生型小鼠视网膜，将每至少10个视网膜混合为一组样本，置于冰预冷的1×磷酸盐缓冲液（PBS）中，立即进行染色质提取、片段化处理，并使用针对Ronin的定制抗体G4275（Dejosez等，2010）、G4275预免疫血清或正常兔IgG（圣克鲁斯生物，货号sc-2027）进行免疫沉淀。随后使用Ion Torrent PGM测序系统对免疫沉淀获取的DNA片段进行测序。实验整体设计：检测出生后第0天（P0）小鼠视网膜基因组中Ronin的结合位点。