Methods and compositions for obtaining marker-free transgenic plants
AssigneeMonsanto Technology LLC
InventorsXudong Ye, Michael W. Petersen, Larry Gilbertson, David Walters, Susan Johnson, Shihshieh Huang, Paul S. Chomet
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Abstract
The invention provides methods and compositions for identifying transgenic seed that contain a transgene of interest, but lack a marker gene. Use of an identification sequence that results in a detectable phenotype increases the efficiency of screening for seed and plants in which transgene sequences not linked to a gene of interest have segregated from the sequence encoding a gene of interest.
Claims46
A method of preparing marker-free seeds from a transgenic plant, comprising the steps of:
- a) obtaining seeds from a transgenic plant, said plant comprising a first DNA segment comprising a nucleic acid of interest and a second DNA segment comprising a marker gene physically linked to a DNA cassette that is operably linked to a promoter functional in said seed, wherein the DNA cassette confers a detectable phenotype comprising altered seed size;
- b) screening the seeds for the absence of the detectable phenotype;
- and
- c) selecting at least a first seed that lacks the detectable phenotype to obtain a seed free of the marker gene.
A DNA construct comprising (a) a first DNA segment comprising left and right T-DNA borders flanking a gene of interest operably linked to a promoter functional in plants, and (b) a second DNA segment comprising a second set of left and right T-DNA borders flanking a promoter functional in a seed operably linked to a DNA cassette that confers a detectable phenotype in seeds that comprise the DNA cassette and a marker gene operably linked to a promoter functional in plants, wherein the detectable phenotype is altered seed size.
A transgenic plant co-transformed with a DNA construct comprising a first DNA segment comprising left and right T-DNA borders flanking a gene of interest operably linked to a promoter functional in plants, and a second DNA construct containing a second DNA segment comprising a second set of left and right T-DNA borders flanking a promoter functional in a seed operably linked to a DNA cassette that confers a detectable phenotype of altered seed size in seeds that comprise the DNA cassette and a marker gene operably linked to a promoter functional in plants.
A DNA construct comprising right and left T-DNA borders, wherein a first DNA segment comprising a gene of interest operably linked to a promoter functional in plants is located downstream in the 5′ to 3′ direction from a right border;
- and a second DNA segment comprising a DNA cassette that confers a detectable phenotype of altered seed size to plant seeds that comprise the DNA cassette and a marker gene operably linked to a promoter functional in plants is located downstream in the 5′ to 3′ direction from a second right border or a left border.
A DNA construct comprising right and left T-DNA borders, wherein a first DNA segment comprising a DNA cassette that confers a detectable phenotype of altered seed size to plant seeds that comprise the DNA cassette and a marker gene operably linked to a promoter functional in plants is located downstream in the 5′ to 3′ direction from a right border, and a second DNA segment comprising a gene of interest operably linked to a promoter functional in plants is located downstream in the 5′ to 3′ direction from a left border.
A DNA construct comprising in the 5′ to 3′ direction first and second right T-DNA borders, wherein a first DNA segment comprising a gene of interest operably linked to a promoter functional in plants is located downstream in the 5′ to 3′ direction from the first right border and a second DNA segment comprising a DNA cassette that confers a detectable phenotype of altered seed size to plant seeds that comprise the DNA cassette and a marker gene operably linked to a promoter functional in plants located downstream in the 5′ to 3′ direction from the second right border.
Dependencies are inferred from the claim text.