|
|
|
| Prokaryotic Expression of Porcine Thyroid Hormone Responsive SPOT14 Gene(THRSP) and Affinity Purification of Its Fused Protein |
|
|
|
|
Abstract Thyroid hormone responsive SPOT14(THRSP), which is a nuclear gene induced by thyroid hormone, expressed predominately in adipose tissue and liver. It is involved in transcriptional regulation of rate-limiting enzymes in lipogenic pathway. It plays an important role on the regulation of animal lipogenesis. In order to further study the function of SPOT14 gene(GenBank accession No. JF951726)， the full cDNA was inserted into the expression vector pET28α(+), and the recombinant expression plasmid pET-28α(+)-S14 was transformed into Escherichia coli BL21(DE3). SDS-PAGE analysis of the IPTG induced expression of the pET-28α(+)-S14 showed that the weight of the target protein was about 20.91 kD, the fusion protein was mainly expressed in the inclusion body, and the best induction condition was 1 mmol/L IPTG for 5 h on 37℃. The fused protein with His-tag was purified by 6xHis Ni-NTA affinity chromatography, and a 21 kD protein was detected by Western blot. This study showed that the recombinant vector pET-28α(+)-S14 can express fused protein in BL21, which paves a way for further study on the gene function and protein function of SPOT14 and the regulation of lipometabolism mechanism
|
|
Received: 27 March 2012
|
|
|
|
| [1] |
ZHU Xiao-Feng, XU Hou-Qiang, CHEN Wei, CHEN Ying-Lian, NI Kai, WU Xiao-Min, NI Meng-Meng, LU Xian-Jun. Cloning, Expression and Bioinformatics Analysis of IGF-1 and IGF-2 Genes in Congjiangxiang Pigs (Sus scrofa)[J]. 农业生物技术学报, 2019, 27(8): 1382-1391. |
|
|
|
|
