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Published on: February 21, 2018
[Expression of the gene encoding human leukocyte differentiation antigen 5D4]
1Department of Immunology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
Summary
This study confirms 5D4 cDNA is full length and describes the expression of 5D4 protein and polypeptides in E. coli, paving the way for monoclonal antibody development.
Area of Science:
- Molecular Biology
- Immunology
- Protein Expression
Background:
- The 5D4 protein's full-length cDNA is crucial for understanding its function and for developing targeted therapeutics.
- Monoclonal antibodies are vital tools for protein research and diagnostics.
Purpose of the Study:
- To verify the integrity of the 5D4 cDNA sequence.
- To produce 5D4 protein fragments as antigens for generating monoclonal antibodies.
Main Methods:
- Construction of recombinant expression vectors (pGEX-5X-3-5D4).
- Expression of 5D4 polypeptides in Escherichia coli DH5 alpha.
- Detection of 5D4 mRNA expression using Northern blot and RT-PCR.
Main Results:
- Northern blot analysis revealed five distinct 5D4 mRNA transcripts (1.9-10.0 kb) with varying tissue distribution.
- A specific 1.9 kb transcript was uniquely detected in spleen and small intestine.
- 5D4 polypeptides were successfully expressed in E. coli.
Conclusions:
- The 1,846 bp 5D4 cDNA sequence is confirmed as full-length.
- Full-length 5D4 protein and a specific N-terminal polypeptide fragment were expressed in E. coli, suitable for antibody production.
