›› 2019, Vol. 39 ›› Issue (1): 4-.doi: 10.3969/j.issn.1674-8115.2019.01.002

• Original article (Basic research) • Previous Articles     Next Articles

Isolation and identification ofOct4 RNA-binding proteins

MA Ji1*, GUO Chuan-liang2, 3*, FAN Shu-yue1, 3, XUE Yan1, 2, 3, ZENG Fan-yi1, 2, 3   

  1. 1. Department of Histoembryology, Genetics & Development, Shanghai Jiao Tong University College of Basic Medical Sciences, Shanghai 200025, China; 2. Shanghai Institute of Medical Genetics, Childrens Hospital of Shanghai Jiao Tong University, Shanghai 200040, China; 3. Shanghai Key Laboratory of Embryo and Reproduction Engineering, National Health Commission of the Peoples Republic of China Key Laboratory of Medical Embryogenesis and Developmental Molecular Biology, Shanghai 200040, China
  • Online:2019-01-28 Published:2019-02-27
  • Supported by:
    China National Basic Research Program, 2014CB964701, 2014CB964703; National Natural Science Foundation of China, 31871484; Constructing Project of Clinical Medical Centers and Clinical Key Disciplines of Shanghai, 2017ZZ02019; Natural Science Foundation of Shanghai, 16ZR1428600

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Abstract: Objective · To identify the RNA binding proteins of Oct4, a key factor of embryonic stem cells. Methods · Total RNA was extracted moembryonic stem cells (R1), which was reverse-transcribed into cDNA. Then PCR product of Oct4 mRNA were transcribed into Oct4 mRNA. Finally the candidate RNA-binding proteins were eluted applying the streptomycin affinity chromatography, and submitted for high-performance liquid chromatography combined with the mass spectrometry. The identified RNA binding proteins were further confirmedRNA immunoprecipitation (RIP). Results · 121 RNA binding proteins of Oct4 3 untranslated region (UTR) and Oct4 5 UTR were identified with liquid chromatography / mass spectrometry. There were 11 proteins with the number of peptide spectrum matches more or equal to 2, and 7 of them were selected for additional confirmation using RIP method. RIP results showed that Oct4 interacted with DSP, SOD1, LMNA, NPM1, PSIP1, NCL and HDGF. Among them, HDGF had the strongest binding ability to Oct4 mRNA. Conclusion · Identification of Oct4 RNA binding proteins will provide a theoretical basis for the regulation mechanism of Oct4, and will be a basis for the further study of its function.

Key words: Oct4, RNA binding protein, embryonic stem cell, RNA immunoprecipitation

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