Retinoblastoma is an infant cancer that results from loss of RB1 expression in both alleles. The RB1 gene was the first reported cancer suppressor gene; however, the mechanism by which RB1 loss causes cancer in the retina has not yet been clarified. Human‐induced pluripotent stem cells (iPSCs) provide an ideal tool for mechanistic research regarding retinoblastoma. However, because RB1 is a tumor suppressor, loss of both alleles of RB1 in human iPS cells may affect the phenotype of the cells. To examine this possibility, we established human iPSCs with deletions in both alleles of RB1 by CRISPR/Cas9 technique to characterize the associated phenotype. We first examined the expression of RB1 transcripts by RT‐qPCR, and RB1 transcripts were expressed in immature hiPSCs and then the expression levels of RB1 transcripts consistently increased during retinal organoid differentiation in human iPSCs. Expression levels of immature markers including SSEA4, OCT3/4 and NANOG were indistinguishable between control iPSCs and RB1 knockout iPSCs. Proliferative activity was also unaffected by homozygous RB1 deletion. Taken together, we showed that homozygous deletion of RB1 did not affect the maturation and proliferation statuses of human iPSCs.

中文翻译：

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人类诱导的多能干细胞携带纯合RB1基因缺失的表征。

视网膜母细胞瘤是由两个等位基因中RB1表达缺失引起的婴儿癌症。该RB1基因是首次报道的抑癌基因; 然而，RB1丢失导致视网膜癌的机制尚未阐明。人类诱导的多能干细胞（iPSC）为有关视网膜母细胞瘤的机理研究提供了理想的工具。但是，由于RB1是肿瘤抑制因子，因此在人iPS细胞中RB1的两个等位基因的缺失可能会影响细胞的表型。为了检查这种可能性，我们建立了人类iPSC，在RB1的两个等位基因中都有缺失通过CRISPR / Cas9技术来表征相关的表型。我们首先通过RT-qPCR检查了RB1转录物的表达，然后在未成熟的hiPSCs中表达了RB1转录物，然后在人iPSC的视网膜类器官分化过程中RB1转录物的表达水平持续增加。在对照iPSC和RB1基因敲除iPSC之间，包括SSEA4，OCT3 / 4和NANOG在内的未成熟标记的表达水平是无法区分的。增殖活性也不受纯合RB1缺失的影响。两者合计，我们表明RB1的纯合删除不影响人类iPSCs的成熟和增殖状态。