Induced pluripotent stem cells are usually derived by reprogramming transcription factors (OSKM), such as octamer-binding transcription factor 4 (OCT4), (sex determining region Y)-box 2 (SOX2), Krüppel-like factor 4 (KLF4), and cellular proto-oncogene (c-Myc). However, the genomic integration of transcription factors risks the insertion of mutations into the genome of the target cells. Recently, the clustered regularly interspaced short palindromic repeat-associated protein 9 (CRISPR/Cas9) system has been used to edit genomes. In this work, dCas9-VPR (dCas9 with a gene activator, VP64-p65-Rta (VPR), fused to its c-terminus) and guide RNA (gRNA) combined to form ribonucleoproteins, which were immobilized on magnetic peptide-imprinted chitosan nanoparticles. These were then used to activate OSKM genes in human embryonic kidney (HEK) 293T cells. Four pairs of gRNAs were used for the binding site recognition to activate the OSKM genes. Transfected HEK293T cells were then prescreened for the high expression of OSKM proteins by immunohistochemistry images. The optimal gRNAs for OSKM expression were identified using quantitative real-time polymerase chain reaction and the staining of OSKM proteins. Finally, we found that the activated expression of one of the OSKM genes is up to three-fold higher than that of the other genes, enabling precise control of the cell differentiation.

诱导多能干细胞通常是通过重新编程转录因子（OSKM）来获得的，例如八聚体结合转录因子4（OCT4），（性别决定区Y）-box 2（SOX2），克鲁勃氏样因子4（KLF4）和细胞原癌基因（c-Myc）。然而，转录因子的基因组整合存在将突变插入靶细胞基因组的风险。最近，簇状规则间隔的短回文重复相关蛋白9（CRISPR / Cas9）系统已用于编辑基因组。在这项工作中，dCas9-VPR（带有基因激活剂VP64-p65-Rta（VPR）的dCas9，融合到其c端）和引导RNA（gRNA）结合形成核糖核蛋白，将其固定在磁性肽印迹壳聚糖上纳米粒子。然后将它们用于激活人胚胎肾脏（HEK）293T细胞中的OSKM基因。使用四对gRNA进行结合位点识别以激活OSKM基因。然后通过免疫组织化学图像对转染的HEK293T细胞进行OSKM蛋白高表达预筛选。使用定量实时聚合酶链反应和OSKM蛋白染色确定了OSKM表达的最佳gRNA。最后，我们发现一个OSKM基因的激活表达比其他基因的激活表达高三倍，从而能够精确控制细胞分化。使用定量实时聚合酶链反应和OSKM蛋白染色确定了OSKM表达的最佳gRNA。最后，我们发现一个OSKM基因的激活表达比其他基因的激活表达高三倍，从而能够精确控制细胞分化。使用定量实时聚合酶链反应和OSKM蛋白染色确定了OSKM表达的最佳gRNA。最后，我们发现一个OSKM基因的激活表达比其他基因的激活表达高三倍，从而能够精确控制细胞分化。