Vectors for plant transformation and methods of use

A method for producing a transgenic plant containing a polynucleotide of interest but being substantially free of non-T-DNA, the method comprising:

• (a) introducing into a plurality of plant cells a T-DNA vector comprising:
• (i) a right border, a left border and a T-DNA sequence therebetween, said T-DNA sequence therebetween lacking a functional cytokinin autonomy gene;
• and
• (ii) a non-T-DNA sequence beyond said left border, said non-T-DNA sequence comprising an Agrobacterium origin of replication and a first visual marker gene, said first visual marker gene being a cytokinin autonomy gene;
• (b) selecting a plant cell which expresses the T-DNA sequence and does not visually manifest abnormal cell proliferation, accelerated or deeper “greening,” delayed senescence, or the formation of “shooty” tumors associated with read-through and expression of cytokinin autonomy gene from the non-T-DNA sequence;
• and
• (c) regenerating a transgenic plant from the selected plant cell.

A method for producing a transgenic plant containing a polynucleotide of interest but being substantially free of non-T-DNA, the method comprising:

• (a) transforming a plurality of plant cells with a T-DNA vector comprising:
• (i) a right border, a left border and a T-DNA sequence therebetween, the T-DNA sequence therebetween lacking a functional cytokinin autonomy gene;
• and
• (ii) a non-T-DNA sequence beyond the left border, the non-T-DNA sequence comprising an Agrobacterium origin of replication, a first visual marker gene, and a second visual marker gene, wherein the first visual marker gene is a cytokinin autonomy gene, and the second visual marker gene encodes a visually detectable fluorescent protein;
• (b) selecting a transformed plant cell and its progeny that do not have visible fluorescence upon illumination with an appropriate light source, wherein fluorescence is associated with read-through and expression of the fluorescent protein gene from the non-T-DNA sequence;
• and
• (c) regenerating a transgenic plant from the selected plant cell.

A method for producing a transgenic plant containing a polynucleotide of interest but being substantially free of non-T-DNA, the method comprising:

• (a) transforming a plurality of plant cells with a T-DNA vector comprising:
• (i) a right border, a left border and a T-DNA sequence therebetween, the T-DNA sequence therebetween containing a polynucleotide of interest and lacking a functional cytokinin autonomy gene;
• and
• (ii) a non-T-DNA sequence beyond the left border, the non-T-DNA sequence comprising an Agrobacterium origin of replication, a first visual marker gene, and a second visual marker gene, wherein the first visual marker gene is a cytokinin autonomy gene, and the second visual marker gene encodes a protein that directs the synthesis of a visually detectable chemical compound;
• (b) selecting a transformed plant cell and its progeny that do not have visible accumulation of the chemical compound, which is associated with read-through and expression of the pigment synthesis gene from the non-T-DNA sequence;
• and
• (c) regenerating a transgenic plant from the selected plant cell.