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Water deficit-inducible promoters

A promoter, inducible technology, applied in organic chemistry, biochemical equipment and methods, sugar derivatives, etc., can solve problems such as adverse development or morphological effects

Inactive Publication Date: 2010-02-17
MENDEL BIOTECHNOLOGY INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These promoter sequences can be used to regulate protein expression during periods of drought or other water-deficient conditions, and thus can be used to induce overexpression of polypeptides that, when desired, confer increased water-deficient tolerance, but No adverse developmental or morphological effects that can be associated with its constitutive overexpression

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment I

[0068] Example I Microarray time course experiment and selection criteria

[0069] Initially, candidate drought-inducible promoters were selected based on the conducted drought time course TxP experiment. In this experiment, clay pots of fully watered late-continuous-leaf Arabidopsis plants grown under short-day conditions were transferred to absorbent paper, and further watering was stopped during the subsequent drought phase. Data are generated for five drought stages determined by physiology: mild stress, moderate stress, severe stress and two stages after rewatering. The stress state of each plant is determined by measuring the physiological indicators of drought, which include relative water content, photosynthetic carbon assimilation, and ABA and proline levels. Leaf tissue samples were collected daily during the two-week drought period, and samples from plants with similar physiological performance for each predetermined state were combined for microarray analysis. The ...

Embodiment II

[0074] Example III Preparation of transgenic plants

[0075] Promoter cloning . For genes showing an appropriate regulation pattern, an upstream sequence of approximately 1.2 kb was cloned by PCR (unless the region contains another gene, in which case cloned up to the upstream sequence of the next gene). Each promoter was cloned into an expression vector (the vector used in this study may include SEQ ID NOs: 10-54, and SEQ ID NOs: 10-27, 33-36, 43-45, and 51- were tested in plants. 54) Before the green fluorescent protein (GFP) or polynucleotide encoding a transcription factor such as SEQ ID NOs: 55, 57, 59, 61 or 59, the transcription factor was shown to provide improved tolerance to water shortage. In some of these cases, the transcription factor constitutively overexpressed also produced harmful morphological effects in the plant.

[0076] Transform . Based on the method of Bechtold and Pelletier (1998) Methods Mol. Biol. 82: 259-266, Arabidopsis thaliana was transformed t...

Embodiment IV

[0082] Example IV GFP fusion expression mode

[0083] Although most of the cloned promoter fragments show the necessary sequences for RNA-driven drought-inducible expression, it is not known whether the elements required for efficient protein translation during stress are included in these constructs. In order to evaluate this problem, a promoter-GFP fusion was used to visually observe the accumulation of GFP protein during and after dehydration treatment. All nine promoters were examined, and in particular it was found that the three promoters prAT1G52690 (from LEA protein), prAT5G52300 (from RD29B) and prAt5G43840 (from heat shock TF G1947) drive high levels of detectable proteins during water stress.

[0084] In multiple events, the promoter from prAT1G52690 (SEQ ID NO: 4) was reliably and strongly induced after drought. The expression level measured by GFP fluorescence is stronger than either the constitutively expressed cauliflower mosaic (CaMV) 35S promoter or the RD29A str...

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Abstract

Water deficit-inducible promoter sequences were identified that may be used to produce transgenic plants that are more tolerant to water deficit and related hyperosmotic stresses than control plants,and yet are wild-type or nearly wild type in appearance. Any of these water deficit-inducible promoters may be incorporated into an expression vector that comprises a polynucleotide regulated by one such promoter and which encodes a polypeptide that, when ectopically expressed, improves water deficit tolerance in plants that are similar to control plants in their morphology and development.

Description

[0001] Joint Research Agreement [0002] In the field of functional genomics and characterization of plant genes for the improvement of plants, the present invention was completed by or on behalf of Mendel Biotechnology, Inc. and Monsanto Corporation as a result of activities carried out within the scope of the following joint research agreement. The joint research agreement was effective on or before the date of the present invention. Invention field [0003] The present invention relates to plant genomics, and more specifically to water deficit inducible promoters that mediate gene expression during the plant's response to water deficit. Background of the invention [0004] In the natural environment, plants often grow under adverse conditions, including water-deficient conditions such as drought, a form of severe low water availability usually characterized by chronic water shortages. Water shortage or dehydration (water deprivation) can delay growth and development, reduce prod...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H1/00C07H21/04C12N15/82C12N5/04
CPCC12N15/8273C12N15/8237
Inventor 彼得·P·里皮第汉斯·E·豪尔坦罗德里克·W·库米摩托奥利弗·J·拉特科力菲
Owner MENDEL BIOTECHNOLOGY INC
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