Genetic engineering of plant chloroplasts
AssigneeAuburn University
InventorsHenry Daniell, Bruce A. McFadden
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Abstract
Novel chimeric constructions and methods for their use are provided for expression of exogenous genes in a plant chloroplast. Particularly, expression is achieved by the use of a chloroplast or bacterial 5′ untranslated region in the expression cassette. The expression cassette may be integrated into the chloroplast genome by the use of chloroplast DNA flanking sequences, or may replicate autonomously if provided with a chloroplast origin of replication. Plants and cells containing the transformed chloroplasts are also provided. The constructs may be used with both monocotyledenous and dicotyledenous chloroplasts.
Claims20
A DNA construct comprising:
- as components, a transcriptional initiation region from a gene capable of expression in a chloroplast joined to a heterologous DNA sequence encoding a polypeptide of interest, wherein transcription of said DNA sequence is regulated by said initiation region, and a transcriptional termination region.
A chloroplast expression vector comprising:
- a transcriptional initiation region from a gene capable of expression in a chloroplast, a DNA sequence comprising at least one cloning site and a transcriptional termination region.
A replication vector comprising:
- a DNA fragment comprising a replication origin capable of providing for autonomous replication in a chloroplast, a transcriptional initiation region from a gene capable of expression in a chloroplast, a DNA sequence comprising at least one cloning site and a transcriptional termination region.
A chloroplast comprising:
- a DNA construct comprising, as components, a transcriptional initiation region from a gene capable of expression in a chloroplast joined to a heterologous DNA sequence encoding a polypeptide of interest, wherein transcription of said DNA sequence is regulated by said initiation region, and a transcriptional termination region, wherein said components are operably linked in vitro.
A chloroplast comprising:
- a chloroplast expression vector comprising, as components, a DNA fragment comprising a replication origin capable of providing for autonomous replication in a chloroplast, a transcriptional initiation region from a gene capable of expression in a chloroplast, a DNA sequence encoding a polypeptide of interest and a transcriptional termination region, wherein said components are operably linked in vitro.
A method for introducing heterologous DNA into a chloroplast, said method comprising:
- transforming a chloroplast in a plant cell with an expression vector comprising, as components, a transcriptional initiation region from a gene capable of expression in a chloroplast, a heterologous DNA sequence encoding a polypeptide of interest and a transcriptional termination region, wherein said components are operably linked in vitro.
A method for specifically modifying the phenotype of a chloroplast distinct from other organelles, said method comprising:
- introducing into a chloroplast in a plant cell, a chloroplast expression vector comprising, as components, a DNA fragment comprising a chloroplast replication origin, a transcriptional initiation region from a chloroplast gene, a DNA sequence encoding a polypeptide of interest and a transcriptional termination region, wherein said components are operably linked in vitro and are functional in said chloroplast;
- and
- growing said cell whereby the phenotype is modified as a result of expression of said DNA sequence.
Dependencies are inferred from the claim text.