Methods and compositions for introduction of exogenous dsRNA into plant cells

Patent metadata

Field	Value
Publication number	US-9433217-B2
Application number	US-201213585947-A
Country	US
Kind code	B2
Filing date	Aug 15, 2012
Priority date	Aug 16, 2011
Publication date	Sep 6, 2016
Grant date	Sep 6, 2016

Abstract

This invention provides a method to silence an endogenous target gene expression in plants by applying a specific dsRNA onto the exterior surface of a plant. Application, such as by spraying or brushing a plant with dsRNA is done without wounding the plant tissue and cells such as by mechanical-type wounding, particle bombardment or mechanical infection with viral vectors. The present invention enables the regulation of gene expression in plants. In some embodiments of the invention, the dsRNA is directed to an essential gene of a plant pathogen or pest, whereby the pathogen and/or pest damage is controlled, resulting in desired agronomic performance.

First claim

What is claimed is:

1. A method of integrating dsRNA into a plant cell to silence an endogenous target gene of a plant comprising:
a) providing a formulation comprising a gene-specific dsRNA, water, and a plant hormone Brassinosteroid, wherein the Brassinosteroid is at 1.0 micromolar, and wherein the gene-specific dsRNA is derived from the endogenous target gene encoding a glutamine synthetase;
b) applying the formulation to the leaf surface of a live plant, wherein the dsRNA is assimilated from the external leaf surface into cells of the plant leaf, wherein the silencing efficiency of the endogenous target gene is increased five to eight fold compared to a formulation containing the dsRNA without the plant hormone Brassinosteroid, and wherein the plant is a maize plant.

2. The method of claim 1 wherein the step of applying the formulation is to a live plant about 12 days from germination.

3. The method of claim 1 wherein the gene-specific dsRNA is derived from the glutamine synthetase cDNA sequence depicted in SEQ ID NO: 2.

4. A method of integrating dsRNA into a plant cell to silence an endogenous target gene of a plant comprising:
a) providing a formulation comprising a gene-specific dsRNA, water, and a plant hormone Brassinosteroid, wherein the dsRNA is at about 250 nanograms per microliter, wherein the Brassinosteroid is at 1.0 micromolar, and wherein the gene specific dsRNA is derived from the endogenous target gene encoding a glutamine synthetase; and
b) applying the formulation to the leaf surface of a live plant, wherein the dsRNA is assimilated from the external leaf surface into cells of the plant leaf, wherein the silencing efficiency of the endogenous target gene is increased five to eight fold compared to a formulation containing the dsRNA without the plant hormone Brassinosteroid, and wherein the plant is a maize plant.

5. The method of claim 4 wherein the step of applying the formulation is to a live plant about 12 days from germination.

6. The method of claim 4 wherein the gene-specific dsRNA is derived from the glutamine synthetase cDNA sequence depicted in SEQ ID NO: 2.

7. A method of integrating dsRNA into a plant cell to silence an endogenous target gene of a plant comprising:
a) providing a formulation comprising a gene-specific dsRNA, water, and a plant hormone Brassinosteroid, wherein the Brassinosteroid is at 1.0 micromolar, and wherein the gene-specific dsRNA is derived from the glutamine synthetase cDNA sequence depicted in SEQ ID NO: 2; and
b) applying the formulation to the leaf surface of a live plant about 12 days from germination, wherein the dsRNA is assimilated from the external leaf surface into cells of the plant leaf, wherein the silencing efficiency of the endogenous target gene is increased five to eight fold compared to a formulation containing the dsRNA without the plant hormone Brassinosteroid, and wherein the plant is a maize plant.

Assignees

• Tang Guo-Qing
• Syngenta Participations Ag

Inventors

• Tang Guo-Qing

Classifications

• C12N15/8218

  Antisense, co-suppression, viral induced gene silencing [VIGS], post-transcriptional induced gene silencing [PTGS] · CPC title

• C12N15/8206

  by physical or chemical, i.e. non-biological, means, e.g. electroporation, PEG mediated · CPC title

• A01N57/16 Primary

  containing heterocyclic radicals · CPC title

• A01N43/22

  rings with more than six members · CPC title

• A01N2300/00

  Combinations or mixtures of active ingredients covered by classes A01N27/00 - A01N65/48 with other active or formulation relevant ingredients, e.g. specific carrier materials or surfactants, covered by classes A01N25/00 - A01N65/48 · CPC title