Codon optimization and ribosome profiling for increasing transgene expression in chloroplasts of higher plants
AssigneeThe Trustees of the University of Pennsylvania
InventorHenry Daniell
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Abstract
Methods for improving transgene in chloroplasts are disclosed along with improved transgenes so produced and methods of use thereof for the treatment of disease. Specifically, the methods comprising analyzing the native sequence of a nucleic acid encoding a protein of interest and replacing codons in said sequence with those preferentially used in psbA genes in chloroplasts in higher plants.
Claims16
A method for increasing translation of a transgene encoding a protein of interest in a chloroplast, said method comprising
- a) analyzing the native sequence of a nucleic acid encoding said protein of interest and replacing codons in said sequence with those preferentially used in psbA genes in chloroplasts in higher plants and optionally performing ribosome profiling and removing any codons that cause stalling of ribosomes during translation;
- b) producing a synthetic, codon optimized sequence and cloning said sequence into a chloroplast transformation vector, said synthetic sequence being operably linked to 5′ and 3′ regulatory elements for suitable for expression in said chloroplast;
- c) transforming a target plant with said vector, under conditions whereby said therapeutic protein is expressed, wherein replacing said codons causes at least a two fold increase in protein expression relative to expression levels observed using the native sequence.
A synthetic VP1 protein encoded by the nucleic acid shown in FIG. 4A.
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A synthetic mutanase enzyme encoded by the nucleic acid shown in FIG. 18 A.
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Dependencies are inferred from the claim text.