Objective To study the function of CTCF during erythroid differentiation and the relevant mechanism. Methods　CTCF was knockdown in K562 cells. Hemin inducing was performed for four days. Quantitative real-time polymerase chain reaction (qRT-PCR) were performed to detect expression of globin genes in control and CTCF knockdown K562 cells, and to evaluate the role of CTCF in erythroid differentiation. Global gene expression pattern after CTCF knockdown was analyzed. Both array data and public data were analyzed to speculate potential regulating mechanism. Chromatin immunoprecipitation (ChIP) was performed to further validate our hypothesis. Results Along with Hemin inducing, expressions of globin genes HBE and HBG both increased in control and CTCF knockdown K562 cells, while expressions of globin genes HBE and HBG in CTCF knockdown K562 cells were lower than that in control cells, indicating CTCF knockdown inhibited expressions of globin genes during erythroid differentiation of K562 cells. 1 128 differentially expressed genes (DEGs) were identified through genome wide gene expression array. Among them, 616 genes were up-regulated in CTCF knockdown cells while 512 genes were down-regulated. IPA analyses showed that the main function items of DEGs included development and function of hematological system. Public data suggested the potential correlation of CTCF-GATA1-ALAS2, and GATA1 was found binding at the ALAS2 gene locus. The ChIP-qPCR results indicated that knockdown CTCF in K562 was able to decrease the binding of GATA1 at ALAS2 gene locus. Conclusion CTCF  plays a role in K562 erythroid differentiation may through regulating interaction between GATA1 and ALAS2 gene locus.