Methods for using artificial polynucleotides and compositions thereof to reduce transgene silencing
The materials and methods disclosed provide for polynucleotide molecules sufficiently divergent from polynucleotides naturally contained in plants, or polynucleotides previously introduced into plants as transgenes to permit trait stacking in plant breeding methods or plant transformation methods. The disclosure also provides for methods and compositions to detect the polynucleotides of the invention in plants.
1. A method to avoid silencing of a transgene comprising SEQ ID NO:16 operably linked to a polynucleotide encoding a chloroplast transit peptide in a transgenic plant, comprising the steps of:
(a) obtaining a DNA construct comprising an artificial polynucleotide encoding a chloroplast transit peptide operably linked to SEQ ID NO:18;
(b) transforming said DNA construct into a plant cell; and
(c) regenerating said plant cell into a fertile transgenic plant,
wherein said fertile transgenic plant comprises both said artificial polynucleotide and said transgene, and wherein silencing of said artificial polynucleotide and said transgene is avoided.
2. An artificial polynucleotide molecule comprising SEQ ID NO:18.
3. A DNA construct comprising: a promoter molecule that functions in plants, operably linked to said artificial polynucleotide molecule of claim 2 .
4. A plant cell, plant or progeny thereof comprising the DNA construct of claim 3 , wherein said DNA construct further comprises a polynucleotide encoding a chloroplast transit peptide operably linked to said artificial polynucleotide molecule.
5. The plant or progeny thereof of claim 4 , wherein said plant is selected from the group consisting of sugarcane, wheat, corn, rice, soybean, cotton, potato, canola, turf grass, forest trees, grain sorghum, vegetable crops, ornamental plants, forage crops, and fruit crops.
6. A plant cell comprising at least two polynucleotides, wherein said two polynucleotides encode the same polypeptide and at least one of the polynucleotides is SEQ ID NO:18 operably linked to a polynucleotide encoding a chloroplast transit peptide.
7. A plant or progeny thereof comprising said plant cell of claim 6 .
8. A plant cell, plant, or progeny thereof comprising an artificial polynucleotide molecule comprising SEQ ID NO:18 operably linked a polynucleotide encoding a chloroplast transit peptide.
9. A DNA detection kit comprising at least one isolated DNA molecule, wherein said isolated DNA molecule is selected from the group consisting of SEQ ID NO:26 and SEQ ID NO:27, wherein said DNA molecule is useful as a DNA probe or DNA primer.
10. A method to avoid transgene silencing in transgenic plants comprising the steps of:
(a) obtaining said plant cell of claim 6 ; and
(b) regenerating said plant cell into a fertile transgenic plant, wherein said fertile transgenic plant comprises both said polynucleotides, and wherein silencing of said polynucleotides is avoided.