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Stress tolerance in plants through selective inhibition of trehalose-6-phosphate phosphatase

A technology for plants and plant cells, applied in plant products, botanical equipment and methods, angiosperms/flowering plants, etc., and can solve problems such as small improvements in drought tolerance

Inactive Publication Date: 2008-01-30
SYNGENTA PARTICIPATIONS AG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Some groups achieved improved trehalose accumulation, and most reported small improvements in drought tolerance despite the overall growth defects observed in the transgenic plants

Method used

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  • Stress tolerance in plants through selective inhibition of trehalose-6-phosphate phosphatase
  • Stress tolerance in plants through selective inhibition of trehalose-6-phosphate phosphatase
  • Stress tolerance in plants through selective inhibition of trehalose-6-phosphate phosphatase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Example 1: Identification and acquisition of ZmTPP1 gene

[0068] The first vascular plant trehalose-6-phosphate phosphatase gene was cloned from Arabidopsis thaliana by complementation of yeast tps2 deletion mutants (Vogel et al. 1998). At that time it was shown that the genes designated AtTPPA and AtTPPB (GenBank accessions AF007778 and AF007779) had trehalose-6-phosphate phosphatase activity. The AtTPPA and AtTTPB protein sequences were used for TBLASTN queries of maize and rice sequence databases. Sequence alignments group hits into individual genes. Figure 2A is a schematic diagram depicting the alignment defining ZmT6PP-1. Three maize T6PP homologues and two rice T6PP homologues were identified. The cDNA sequences corresponding to the predicted protein sequences of each gene - ZmT6PP-1, -2 and -3 and OsT6PP-1 and -2 - are shown in SEQ ID NO. 1, 2, 3, 4 and 5, respectively. Figure 3 shows the overall alignment of these T6PPs with Arabidopsis T6PPs. The relatio...

Embodiment 2

[0072] Embodiment 2: Construction of Rab17 expression cassette

[0073] An expression cassette based on the maize Rabl7 gene has been found to be drought-induced in vegetative tissues (Vilardell et al., 1990) and develops expression in maturing seeds (Vilardell et al., 1991). One embodiment of the present invention is to provide a nucleic acid construct comprising a drought-inducible promoter in vegetative tissue operably linked to a nucleic acid molecule, wherein when the nucleic acid molecule is in a plant cell When expressed, it reduces the expression of the endogenous T6PP gene in plant cells.

[0074] The present invention includes a nucleic acid construct having a promoter derived from the 5' region of Rab17 gene and exhibiting promoter activity in plants.

[0075] The present invention also includes a nucleic acid construct comprising the entire or partial sequence of the nucleic acid encoding T6PP-RNAi, wherein the Rab17 promoter drives the T6PP-RNAi expression casset...

Embodiment 3

[0087] Embodiment 3: Construction of T6PP-RNAi expression cassette

[0088] Figure 6 shows the primers used to generate the T6PP-RNAi gene. Two PCR fragments were generated from the pCR-4-TOPO-ZmT6PP-NS template (Figure 7). Fragment 1 (SEQ ID NO. 15) contains part of the CMVpSPORT6 vector as a loop in the T6PP-RNAi gene product. Fragment 1 was amplified from pCR-4-TOPO-ZmT6PP-1 using high-fidelity PCR in a 50 μL reaction mixture consisting of: 1 μL miniprep pCR-4-TOPO-ZmT6PP-NS DNA, 200 μM dNTPs, 20 μM oligonucleotides Nucleotide primer 001L (5'-ATAGGCGCGCCATGTTGGAGATGACAGAACAGATC-3') (SEQ ID NO.38), 20 μM oligonucleotide primer 002R (5'-ATACCGCGGGGACTGTCCTGCAGGTTTAAACG-3') (SEQ ID NO.39), 5 μL 10 × clone Pfu buffer and 2.5 units of Pfuturbo DNA polymerase (Stratagene, Cat. No. 600252) in a final volume of 50 μL. The thermal cycling program was at 95°C for 30 seconds, followed by 40 cycles (95°C for 10 seconds, 65°C for 60 seconds, 72°C for 2 minutes), followed by 72°C for ...

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Abstract

The present invention relates to transgenic plants comprising an isolated DNA molecule comprising a polynucleotide that encodes a nucleic acid that down-regulates an endogenous T6PP gene, wherein the polynucleotide is under the control of a promoter that is stress-inducible and is expressed predominantly in vegetative tissue. The promoter may also be developmentally expressed in maturing kernels. Expression of the polynucleotide results in the increased availability of carbon to developing florets / kernels when plants are subject to environmental stress, such as a water deficit. The DNA molecule of the invention thereby permits more photosynthate to be directed to the developing ovules / embryos resulting in stabilized yield in growing environments that are subject to periodic stress.

Description

technical field [0001] The present invention includes stress-responsive expression of nucleic acid sequences capable of down-regulating trehalose-6-phosphate phosphatase activity, so as to increase plant yield and / or enhance plant abiotic stress tolerance. Background technique [0002] Abiotic stresses affect plant development in different ways, depending on the timing, severity and duration of the stress. For example, maize plants are relatively drought tolerant and can tolerate moderate to severe drought early and late in the growing season. However, maize is quite sensitive to water stress during the 10-14 days before and after flowering. Drought tolerant corn grown in US corn regions typically experiences water stress during the summer season during flowering. This stress usually takes the form of slowed kernel formation due to ovule / embryo abortion. Briefly, when roots experience osmotic stress, they produce abscisis acid (ABA) and transfer it throughout the plant. ...

Claims

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Application Information

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IPC IPC(8): A01H5/00A01H5/10C12N15/82
CPCC12N15/8273C12N15/8237C12N9/16
Inventor M·L·拉格里米尼M·努奇奥N·斯普林格
Owner SYNGENTA PARTICIPATIONS AG
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