Evaluation of novel control elements by construction of eukaryotic expression vectors.
A novel mammalian eukaryotic expression vector for the production of immunoglobulin heavy chain (IgH) genes has been designed. This expression vector contains the variable heavy chain (VH) promoter, the IgH intron enhancer (muE) and the IgH 3' enhancer (3'E). This construct, designated pTIF-1, was stably transfected into the myeloma cell line J558L. A fivefold increase in the expression level of a rearranged IgH gene was observed when using the pTIF-1 vector containing the 3'E compared to an expression vector lacking this enhancer. Interestingly, this positive effect on the expression level of the 3' enhancer appears to be position independent. The introduction of two recently identified Ig control elements, HS3 and HS4, to the vector cassette did not further elevate the expression level in the cell line tested. The pTIF-1 vector can be used for expression of any antibody specificity, using PCR amplification of the VDJ region of interest. Furthermore, the constant region can easily be exchanged, which further facilitates studies to dissect different effector functions of IgH constant genes.
Furebring
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C
Department of Immunotechnology, Lund University, Sweden.
Ohlin
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M
Pettersson
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Borrebaeck
C A
CA
D000818:Animals; D004742:Enhancer Elements, Genetic; D005057:Eukaryotic Cells; D005069:Evaluation Studies as Topic; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics; D051379:Mice; D012045:Regulatory Sequences, Nucleic Acid; D014407:Tumor Cells, Cultured
D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region