Optimizing expression of LZ-8 protein and primary assaying of its immunomodulatory activity
To obtain sufficient recombinant LZ-8 protein (rLZ-8) for assaying its immunosuppressive activity, 12 overlapping oligonucleotides optimized by the usage of the favorite codons in E. coli were synthesized and connected to form a new LZ-8 gene by PCR. The recombinant plasmid pET-28b_lz8 containing th...
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| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Zhejiang University Press
2008-01-01
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| Series: | 浙江大学学报. 农业与生命科学版 |
| Subjects: | |
| Online Access: | https://www.academax.com/doi/10.3785/j.issn.1008-9209.2008.01.002 |
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| Summary: | To obtain sufficient recombinant LZ-8 protein (rLZ-8) for assaying its immunosuppressive activity, 12 overlapping oligonucleotides optimized by the usage of the favorite codons in E. coli were synthesized and connected to form a new LZ-8 gene by PCR. The recombinant plasmid pET-28b_lz8 containing the new LZ-8 gene was constructed by inserting the new gene into the expression plasmid pET-28b. The rLZ-8 protein was induced expression and purified in E. coli BL21. High yield (about 70 mg·L<sup>-1</sup> of induced culture) and purity (about 90%) of rLZ-8 protein were achieved by one-step nickel-affinity chromatography and then assayed by using skin transplantation model in mice. The results show that rLZ-8 protein can be highly expressed after codon usage optimization in E. coli, and the immunological activity of mice injected with the purified rLZ-8 was suppressed. But this immunosuppressive activity of rLZ-8 need to be further evaluated. |
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| ISSN: | 1008-9209 2097-5155 |