Feeder-free derivation of human induced pluripotent stem cells with messenger RNA.

Feeder-free derivation of human induced pluripotent stem cells with messenger RNA.
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DOI:
10.1038/srep00657
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发表时间:
2012
期刊:
影响因子:
4.6
通讯作者:
Guo, Xirong
Guo, Xirong
中科院分区:
综合性期刊3区
文献类型:
--
作者:
Warren, Luigi;Ni, Yuhui;Wang, Jiwu;Guo, Xirong

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诱导多能干细胞(iPSC)的治疗前景促使人们在将体细胞重编程为多能性时努力规避基因组改变。基于附加型DNA、仙台病毒和信使RNA(mRNA)的方法可以产生“无足迹”的iPSC,其效率等于或超过整合病毒载体所获得的效率。mRNA方法独特地提供了对重编程因子(RF)表达的前所未有的控制,同时避免了清除载体残留痕迹的清除阶段。目前,基于mRNA的重编程是相对费力的,因为需要每天重复约2周以诱导多能性,并且需要使用饲养细胞,这增加了程序的复杂性和可变性,同时引入了非人源生物材料污染的途径。我们通过逐步优化RF鸡尾酒来加速mRNA重编程过程,并利用这些动力学增益来建立一种无饲养层、无异种的方案,该方案削减了iPSC衍生所涉及的时间、成本和精力。
The therapeutic promise of induced pluripotent stem cells (iPSCs) has spurred efforts to circumvent genome alteration when reprogramming somatic cells to pluripotency. Approaches based on episomal DNA, Sendai virus, and messenger RNA (mRNA) can generate “footprint-free” iPSCs with efficiencies equaling or surpassing those attained with integrating viral vectors. The mRNA method uniquely affords unprecedented control over reprogramming factor (RF) expression while obviating a cleanup phase to purge residual traces of vector. Currently, mRNA-based reprogramming is relatively laborious due to the need to transfect daily for ~2 weeks to induce pluripotency, and requires the use of feeder cells that add complexity and variability to the procedure while introducing a route for contamination with non-human-derived biological material. We accelerated the mRNA reprogramming process through stepwise optimization of the RF cocktail and leveraged these kinetic gains to establish a feeder-free, xeno-free protocol which slashes the time, cost and effort involved in iPSC derivation.
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