Methods and compositions for generating dominant short stature alleles using genome editing

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification comprising a deletion of at least a portion of the intergenic region between the endogenous GA20 oxidase_5 and Zm.SAMT genes, and wherein the mutant allele produces a RNA molecule comprising an antisense sequence complementary to all or part of the sense strand of the endogenous GA20 oxidase_5 gene.

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification comprising a deletion of at least a portion of one or more of the following:

• 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any portion thereof, and the 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any portion thereof, of the endogenous Zm.SAMT gene.

The modified corn plant, or plant part thereof, of any one of claims 21-25, wherein the hybridization can cause suppression of a wild-type or mutant allele of the endogenous GA20 oxidase_3 gene, a wild-type or mutant allele of the endogenous GA20 oxidase_5 gene, or a wild-type or mutant allele of both genes.

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification which results in the transcription of an antisense strand of at least an exon, an intron, or an untranslated region (UTR) of the endogenous GA20 oxidase_5 gene, or any portion thereof.

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises the Zm.SAMT gene promoter, or a functional part thereof, operably linked to at least one transcribable antisense sequence of at least an exon, intron or untranslated region (UTR) of the endogenous GA20 oxidase_5 gene, or any portion thereof.

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a sequence selected from the group consisting of SEQ ID NOs:

• 87-105.

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a first sequence and a second sequence;

• wherein the first sequence comprises one or more of the 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.GA20 oxidase_5 gene;
• and wherein the second sequence comprises one or more of the 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.SAMT gene;
• wherein the first sequence and the second sequence are contiguous or separated only by an intervening sequence of fewer than 555 nucleotides.

A modified corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genomic deletion relative to a wild type allele of the endogenous GA20 oxidase_5 locus, wherein the genomic deletion is flanked by a first sequence and a second sequence;

• wherein the first sequence comprises one or more of the 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.GA20 oxidase_5 gene;
• and wherein the second sequence comprises one or more of the 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.SAMT gene.

A method for producing a modified corn plant comprising a mutant allele of the endogenous GA20 oxidase_5 locus, the method comprising:

• a. generating two double-stranded breaks (DSB) in or near the endogenous GA20 oxidase_5 locus in a corn cell using a targeted editing technique;
• b. developing or regenerating from the corn cell a corn plant, or plant part thereof, comprising a mutant allele of the endogenous GA20 oxidase_5 locus.

A method for producing a modified corn plant comprising a mutant allele of the endogenous GA20 oxidase_5 locus, the method comprising:

• a. generating a first and a second double-stranded breaks (DSB) in a corn cell using a targeted editing technique, wherein the first DSB is in a region selected from the group consisting of 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any portion of the foregoing, of the endogenous GA20 oxidase_5 locus, and the intergenic region between the endogenous Zm.GA20 oxidase_5 gene and the endogenous Zm.SAMT gene;
• wherein the second DSB is in a region selected from the group consisting of 5′ UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any portion of the foregoing, of the endogenous Zm.SAMT locus, and the intergenic region between the endogenous Zm.GA20 oxidase_5 gene and the endogenous Zm.SAMT gene;
• b. developing or regenerating from the corn cell a corn plant, or plant part thereof, comprising a genomic deletion, wherein the genomic deletion is flanked by the first DSB and the second DSB.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification comprising a deletion of at least a portion of the transcription termination sequence of the endogenous Zm.SAMT gene, and wherein the mutant allele produces a RNA molecule comprising an antisense sequence complementary to all or part of the sense strand of the endogenous GA20 oxidase_5 gene.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification comprising a deletion of at least a portion of the intergenic region between the endogenous GA20 oxidase_5 and Zm.SAMT genes, and wherein the mutant allele produces a RNA molecule comprising an antisense sequence complementary to all or part of the sense strand of the endogenous GA20 oxidase_5 gene.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification comprising a deletion of at least a portion of one or more of the following:

• 5′UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any portion thereof, and the 5′UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any portion thereof, of the endogenous Zm.SAMT gene.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genome modification which results in the transcription of an antisense strand of at least an exon, an intron, or an untranslated region (UTR) of the endogenous GA20 oxidase_5 gene, or any portion thereof.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises the Zm.SAMT gene promoter, or a functional part thereof, operably linked to at least one transcribable antisense sequence of at least an exon, intron or untranslated region (UTR) of the endogenous GA20 oxidase_5 gene, or any portion thereof.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a sequence selected from the group consisting of SEQ ID NOs:

• 87-105.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a first sequence and a second sequence;

• wherein the first sequence comprises one or more of the 5′UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.GA20 oxidase_5 gene;
• and wherein the second sequence comprises one or more of the 5′UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.SAMT gene;
• wherein the first sequence and the second sequence are contiguous or separated only by an intervening sequence of fewer than 555 nucleotides.

A modified corn plant part, corn cell, or corn tissue comprising a mutant allele of the endogenous GA20 oxidase_5 locus, wherein the mutant allele comprises a genomic deletion relative to a wild type allele of the endogenous GA20 oxidase_5 locus, wherein the genomic deletion is flanked by a first sequence and a second sequence;

• wherein the first sequence comprises one or more of the 5′UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.GA20 oxidase_5 gene;
• and wherein the second sequence comprises one or more of the 5′UTR, 1st exon, 1st intron, 2nd exon, 2nd intron, 3rd exon, 3rd intron, 4th exon, 4th intron, 5th exon, 5th intron, 6th exon, 6th intron, 7th exon, 7th intron, 8th exon, 3′ UTR, and any complementary sequence thereof, and any portion of the foregoing, of the endogenous Zm.SAMT gene.