Molecular cloning and preliminary functional study of TRBP].

Construct prokaryotic expression vector carrying mouse TRBP (TAR RNA-binding protein) gene and test the double-stranded RNA binding ability of TRBP. RT-PCR was used to obtain TRBP cDNA from mouse genomic DNA. Then, we built the His-tag fusion expression vector of TRBP and transformed it into E.coli BL21(DE3). Ni-NTA beads were used to isolate and purify the recombinant protein and vitro transcription was used to get Pre-miR-122. Finally, SDS-PAGE and ITC (isothermal titration calorimetry) assay were both used to validate TRBP's binding ability with Pre-miR-122. We purified the recombinant protein TRBP whose molecular weight is 32.4 kDa. The purified bioactive TRBP protein binding on NI-NTA beads showed that it had a strong binding capacity on Pre-miR-122. We constructed TRBP prokaryotic expression system successfully and studied the double-stranded RNA binding ability of TRBP preliminarily.

Citation

Zhan-jun Liu, Ying Zhu, Wen-gong Yu, Xin-zhi Lu. Molecular cloning and preliminary functional study of TRBP]. Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology. 2012 Feb;28(2):124-6

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PMID: 22304766

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