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US 6,649,409 B1Granted
Method for producing a nucleotide sequence construct with optimized codons for an HIV genetic vaccine based on a primary, early HIV isolate and synthetic envelope BX08 constructs
AssigneeStatens Serum Institut
InventorAnders Fomsgaard
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Abstract
The present invention relates to a method for producing a nucleotide sequence construct with optimized codons for an HIV genetic vaccine based on a primary, early HIV isolate. Specific such nucleotide sequence construct are the synthetic envelope BX08 constructs. The invention further relates to the medical use of such constructs for the treatment and prophylaxis of HIV through DNA vaccine and for diagnostics.
Claims48
A method for producing a nucleotide sequence construct comprising the following steps:
- a) obtaining a first nucleotide sequence of an HIV gene from a patient within the first 12 months of infection;
- b) designing a second nucleotide sequence utilising the most frequent codons from mammalian highly expressed proteins to encode the same amino acid sequence as the first nucleotide sequence of a) encodes
- c) redesigning the second nucleotide sequence of b) so that restriction enzyme sites surrounds the regions of the nucleotide sequence which encode functional regions of the amino acid sequence and so that selected restriction enzyme sites are removed thereby obtaining a third nucleotide sequence encoding the same amino acid sequence as the first and the second nucleotide sequence of a) and b) encode;
- d) redesigning the third nucleotide sequence of c) so that the terminal snuts contain convenient restriction enzyme sites for cloning into an expression vehicle;
- e) producing the snuts between restriction enzyme sites of c) and terminal snuts of d).
2 more elements
A nucleotide sequence construct in isolated form which has a nucleotide sequence with the general formula (I), (II), (III), or (IV) or subsequences thereof
- P1-S495ClaI-S650-720EcoRI-P2-S1265gp120 (I)
- P1-S495ClaI-S650-720EcoRI-P2-S1265XhoI-S1465PstI-P4gp140 (II)
- P1-S495ClaI-S650-720EcoRI-P2-S1265XhoI-S1465PstI-P4gp150 (III)
- P1-S495ClaI-S650-720EcoRI-P2-S1265XhoI-S1465PstI-P4gp160-S2060SacII-P5 (IV)
- wherein P1 designates the nucleotide sequence SEQ ID NO:41, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 90% thereto;
- wherein S495ClaI designates the nucleotide sequence SEQ ID NO:
- 7, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 95%thereto;
- wherein S650-720EcoRI designates the nucleotide sequence SEQ ID NO:
- 9, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 95% thereto;
- wherein P2 designates the nucleotide sequence SEQ ID NO:
- 43, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 85% thereto;
- wherein S1265gp120 designates the nucleotide sequence SEQ ID NO:
- 19, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 70% thereto;
- wherein S1265XhoI designates the nucleotide sequence SEQ ID NO:
- 17, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 80% thereto;
- wherein S1465Psil designates the nucleotide sequence SEQ ID NO:
- 23, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 90% thereto;
- wherein P4gp140 designates the nucleotide sequence SEQ ID NO:
- 57, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 85% thereto;
- wherein P4gp150 designates the nucleotide sequence SEQ ID NO:
- 55, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 85% thereto;
- wherein P4gp160 designates the nucleotide sequence SEQ ID NO:
- 53, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 85% thereto;
- wherein S2060sacII designates the nucleotide sequence SEQ ID NO:
- 33, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 98% thereto;
- and
- wherein P5 designates the nucleotide sequence SEQ ID NO:
- 59, a nucleotide sequence complementary thereto, or a nucleotide sequence with a sequence identity of at least 85% thereto.
25 more elements
Dependencies are inferred from the claim text.