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Published on: April 20, 2018
[Molecular cloning and preliminary functional study of TRBP]
Zhan-jun Liu1, Ying Zhu, Wen-gong Yu
1School of Medicine and Pharmacy, Ocean University of China, Qingdao, China. lzj2385759@126.com
Summary
Researchers successfully created a prokaryotic expression system for mouse TAR RNA-binding protein (TRBP). This system confirmed TRBP
Area of Science:
- Molecular Biology
- Protein Expression
- RNA-Binding Proteins
Background:
- TAR RNA-binding protein (TRBP) plays a crucial role in RNA processing and antiviral defense.
- Understanding TRBP's function requires efficient methods for its expression and purification.
Purpose of the Study:
- To construct a prokaryotic expression system for mouse TAR RNA-binding protein (TRBP).
- To investigate the double-stranded RNA binding ability of the expressed TRBP protein.
Main Methods:
- Mouse TRBP cDNA was amplified using RT-PCR.
- A His-tag fusion expression vector was constructed and transformed into E. coli BL21(DE3).
- Recombinant TRBP was purified using Ni-NTA affinity chromatography, and its binding to Pre-miR-122 was assessed using SDS-PAGE and Isothermal Titration Calorimetry (ITC).
Main Results:
- The recombinant mouse TRBP protein (32.4 kDa) was successfully purified.
- The purified TRBP demonstrated significant binding capacity to Pre-miR-122, a double-stranded RNA molecule.
Conclusions:
- A functional prokaryotic expression system for mouse TRBP was established.
- The study provides preliminary evidence of TRBP's double-stranded RNA binding capability.
