Pioneer Hi-Bred International, Inc.

Boyce Thompson Institute for Plant Research

Other

Gregory D May

Peter B Kipp

Christopher L Baszczynski

Pramod B Mahajan

Tong Zhu

Charles Joel Arntzen

Yue Yun

Ramesh Kumar

Mary Beatty

Gina Marie Zastrow-Hayes

Stephane Deschamps

Nicholas Doane Chilcoat

Stephanie K Clendennen

Kevin Day

Wayne Matthew Crismani

Compositions and methods of identifying and characterizing potential gene editing on-target and off-target sites and/or edits in a nucleic acid are provided.

Methods of identifying and characterizing gene editing variations in nucleic acids

The invention concerns the use of duplex oligonucleotides about 25 to 30 base pairs to introduce site specific genetic alterations in plant cells. The oligonucleotides can be delivered by mechanical (biolistic) systems or by electrpoporation of plant protoplasts. Thereafter plants having the genetic alteration can be generated from the altered cells. In specific embodiments the invention concerns alteration in the gene that encode acid invertase, UDP-glucose pyrophosphorylase, polyphenol oxidase, O-methyl transferase, cinnamyl alcohol dehydrogenase, ACC synthase and ACC oxidase or etr-1 or a homolog of etr-1, and plants having isolated point mutations in such genes.

Use of mixed duplex oligonucleotides to effect localized genetic changes in plants

The invention relates to isolated nucleic acid molecules encoding MutS homologues (MSHs). Such MSH proteins are involved in DNA mismatch-repair processes in organisms. The invention provides isolated nucleic acid molecules comprising MSH2 nucleotide sequences which encode MSH2 proteins and MSH2 nucleotide sequences which encode dominant-negative MSH2 variants. Such MSH2 nucleotide sequences find use in altering mismatch repair, mutation rates and recombination frequencies in both eukaryotic and prokaryotic organisms. The invention also provides isolated nucleic acid molecules comprising MSH2 promoter nucleotide sequences. Such MSH2 promoter nucleotide sequences find use in regulating the expression of genes of interest in plants. Additionally provided are isolated proteins, transformed host cells, and transformed plants, tissues, cells and seeds thereof.

Plant MSH2 sequences and methods of use

The present invention provides isolated and purified genes which are differentially expressed during banana fruit development, and the protein products of these genes. The present invention further provides DNA regulatory elements which are differentially expressed during banana fruit development, chimeric genes comprising these DNA regulatory elements operably linked to heterologous DNA molecules, and plants transformed with said chimeric genes, providing for controlled expression of said heterologous DNA molecules during the development and ripening of the fruit of said plants, or in response to exogenous ethylene signals in said plants. The present invention also provides a method for expression of a heterologous protein in fruit comprising transforming fruiting plants with one or more chimeric genes according to the present invention, exposing said fruit to an endogenous or exogenous ethylene signal, and harvesting fruit containing said heterologous protein. The method of the present invention may further comprise isolated the proteins produced by said method from the harvested fruit. In a particularly preferred embodiment, the heterologous protein is a therapeutic protein, which may be isolated from the harvested fruit, or consumed directly in the transformed fruit by a patient in need of said therapeutic protein.

Banana DNA associated with fruit development

Growing community of inventors

Ardmore, OK, United States of America

Gregory D May