Fruit-specific promoters
Inventors
Thomson, James G. • Thilmony, Roger L. • DASGUPTA, Kasturi • Dardick, Christopher D.
Assignees
US Department of Agriculture USDA
Publication Number
US-11739341-B2
Publication Date
2023-08-29
Expiration Date
2039-05-23
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Abstract
The present disclosure provides genetic constructs containing a promotor that is useful in driving fruit-specific expression in plants. Further provided are expression vectors, transgenic plants, and plant parts containing such genetic constructs, as well as uses thereof.
Core Innovation
The invention relates to genetic constructs containing promoters that drive fruit-specific expression in plants. These promoters comprise sequences selected from SEQ ID NOs: 1-5 or sequences having at least 90% identity thereto. The genetic constructs are operably linked to heterologous nucleotide sequences encoding products of interest, such as RNA molecules or polypeptides involved in various metabolic pathways. The disclosure also includes expression vectors, transgenic plants and plant parts containing these genetic constructs, and methods for using them in fruit breeding to modify fruit phenotypes.
The problem addressed is the limitation of existing fruit-specific promoters available for genetic engineering in many fruit species, which hampers genetic improvement of fruits for traits like growth, ripening, nutritional quality, and shelf life. Traditional breeding methods face challenges such as large plant size, long juvenile phases, and limited genetic gains. Therefore, a need exists for novel promoters providing robust and reliable fruit-specific expression to enable targeted expression of transgenes with minimal adverse effects on normal plant growth and development.
Claims Coverage
The claims focus on a set of genetic constructs, expression vectors, transgenic plants, plant parts, and methods for modifying fruit phenotypes, highlighting several inventive features related to promoters, products of interest, and applications in plants.
Genetic construct with specific fruit-specific promoters
A genetic construct comprising a promoter operably linked to a heterologous polynucleotide encoding a product of interest, where the promoter comprises a sequence as set forth in SEQ ID NO: 2, 3, 4, or 5, or a sequence having an ATCT-motif, a G-Box, an HSE element, an MBS, a CAAT box, and a TATA box, and at least 96% identity to SEQ ID NO: 2, 3, 4, or 5.
Product of interest encoded by the heterologous nucleotide sequence
The product of interest encoded by the heterologous polynucleotide may be an RNA molecule or a polypeptide.
Product of interest involved in metabolic pathways
The product of interest can be involved in metabolic pathways selected from anthocyanin, tocopherol, fatty acid, carotenoid, lycopene, betalain, and flavonoid metabolic pathways.
Product of interest being specific polypeptides
The product of interest may be a polypeptide selected from the group consisting of MYB transcription factor, phytoene synthase (PSY), lycopene cyclase (LCY), and DXP synthase (DXS).
Expression vector comprising the genetic construct
An expression vector comprising the genetic construct that contains the specific promoter operably linked to the product of interest.
Transgenic plant comprising the genetic construct
A transgenic plant comprising the genetic construct with the specified promoter and product of interest.
Plant parts containing the genetic construct
A plant part derived from the transgenic plant that contains the genetic construct, where the plant part may be a stem, branch, root, leaf, flower, fruit, seed, cutting, bud, cell, or portion thereof.
Method for modifying fruit phenotype
A method for modifying a fruit phenotype in a plant, involving transforming a plant cell with the genetic construct, regenerating a plant from the transformed cell, and growing the regenerated plant to produce fruit with the desired modified phenotype.
Specified fruit phenotypes
The fruit phenotype modified by the method may be selected from size, weight, color, shape, firmness, glossiness, flavor, aroma, secondary metabolite content, peel thickness, seed number, juice quality, juice sugar content, juice acid content, juice taste, juice color, and juice yield.
Fruit phenotypes related to secondary metabolite content
Fruit phenotypes may include anthocyanin content, tocopherol content, fatty acid content, carotenoid content, lycopene content, betalain content, and flavonoid content.
Applicable plants
The method and genetic constructs are applicable to plants selected from orange (Citrus sinensis), mandarin (Citrus reticulata), lime (Citrus aurantifolia), grapefruit (Citrus paradisi), lemon (Citrus limon), pomelo (Citrus maxima), citron (Citrus medica), papeda (Citrus micrantha), and Prunus species.
The claims cover genetic constructs containing specified fruit-specific promoters operably linked to heterologous nucleotide sequences encoding products of interest, expression vectors and transgenic plants containing these constructs, plant parts harboring the constructs, and methods of modifying fruit phenotypes by transformation and regeneration of plants with altered fruit traits, applicable to a range of citrus and Prunus species.
Stated Advantages
The promoters enable robust and reliable fruit-specific expression, minimizing adverse effects on plant growth and development.
The use of fruit-specific promoters facilitates targeted genetic modification of fruit traits such as growth, ripening, nutritional quality, and post-harvest shelf life.
The genetic constructs allow modification of secondary metabolite content in fruits, including anthocyanins, tocopherols, carotenoids, lycopene, betalains, and flavonoids.
The method generates seedless tomato fruit without compromising fruit quality or vegetative and reproductive development.
Documented Applications
Engineering fruit traits of agronomic importance such as improved growth, ripening, nutritional quality, and post-harvest shelf life.
Modifying fruit phenotypes including size, weight, color, shape, firmness, glossiness, flavor, aroma, secondary metabolite content, peel thickness, seed number, juice qualities, and yield.
Increasing accumulation of secondary metabolites in fruits such as anthocyanins, tocopherols, fatty acids, carotenoids, lycopene, betalains, and flavonoids for enhanced nutraceutical value.
Creating seedless tomato fruits through transformation with a genetic construct comprising the promoter of SEQ ID NO: 4 or sequences with at least 90% identity thereto.
Use in fruit breeding via transformation, plant regeneration, and growing of transgenic plants exhibiting modified fruit characteristics.
Use in various fruit species including citrus varieties (orange, mandarin, lime, grapefruit, lemon, pomelo, citron, papeda) and Prunus species.
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