Heterologous intron within an immunoglobulin domain
AssigneeBoehringer Ingelheim International GmbH
InventorBarbara ENENKEL
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Abstract
The invention concerns the field of recombinant gene engineering. It concerns novel introns and compositions comprising such introns as well as a method to improve expression of polypeptides from nucleic acids such as cloned genes with heterologous introns, especially genes encoding antibodies and antibody derived fragments, and the production of various polypeptides in eukaryotic host cells using said novel intron sequences as heterologous introns.
Claims23
A transcription unit or expression vector comprising a gene of interest encoding at least one domain of an immunoglobulin gene, comprising at least one heterologous intron located within an immunoglobulin exon, whereby the 5′ and 3′ ends of said exon are defined as occurring in a corresponding native immunoglobulin gene.
A method of producing a protein of interest encoded by a polynucleotide sequence comprising at least one domain of an immunoglobulin gene comprising:
- a) introducing at least one heterologous intron sequence into said domain by placing the intron into a nucleotide sequence of an immunoglobulin exon, whereby the 5′ and 3′ ends of the exon are defined as occurring in a corresponding native immunoglobulin gene, and
- b) introducing the immunoglobulin domain comprising/containing the nucleic acid sequence of step a) into a transcription unit or a expression vector, and
- c) transfecting a cell with said transcription unit or said vector of step b), and
- d) cultivating said cell of step c) under conditions which allow expression of said protein of interest.
A kit comprising
- a vector including at least one intron sequence flanked by suitable recognition sites for restriction enzymes allowing the integration of a first and/or a second nucleotide sequence coding for at least a part of an immunoglobulin gene, and
- instructions and optionally a vector map, and
- optionally a host cell, and
- optionally a cultivation medium for the cultivation of a host cell, and/or
- optionally a selection medium for selecting and cultivating a transfected host cell.
A kit comprising
- a) a vector including at least parts of a transcription unit comprising a first nucleotide sequence comprising at least one heterologous intron located within an immunoglobulin exon or parts of an immunoglobulin exon, whereby the 5′ and 3′ ends of said exon are defined as occurring in a corresponding native immunoglobulin gene sequence, and whereby the nucleotide sequence of this transcription unit comprises suitable recognition sites for restriction enzymes allowing the integration of a second nucleotide sequence coding for another part of the gene of interest, whereby the two nucleotide sequences are operatively linked, and
- b) instructions and optionally a vector map, and
- c) optionally a host cell, and
- d) optionally a cultivation medium for the cultivation of a host cell, and/or
- e) optionally a selection medium for selecting and cultivating a transfected host cell.
Dependencies are inferred from the claim text.