UCE: A uracil excision (USER™)-based toolbox for transformation of cereals

Hebelstrup, Kim H.; Christiansen, Michael W.; Carciofi, Massimiliano; Tauris, Birgitte; Brinch-Pedersen, Henrik; Holm, Preben Bach

doi:10.1186/1746-4811-6-15

Cited by 22 publications

(27 citation statements)

References 23 publications

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“…Stable transgenic rice lines were generated by Agrobacterium -mediated transformation using pUCE D-Hord:TP-StGWD:NOS vector (Supplementary Fig. 1) that enables endosperm-specific overexpression of transgenic protein and targeting to the amyloplasts by an N-terminal transit peptide (TP) 25 . Sixteen transgenic lines among 24 candidate plants in the T 0 generation were identified by genomic PCR for hygromycin phosphotransferase ( HPT ) and StGWD1 as selection markers (Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

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Highly phosphorylated functionalized rice starch produced by transgenic rice expressing the potato GWD1 gene

Chen

Sun

Zhou

et al. 2017

Sci Rep

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Starch phosphorylation occurs naturally during starch metabolism in the plant and is catalysed by glucan water dikinases (GWD1) and phosphoglucan water dikinase/glucan water dikinase 3 (PWD/GWD3). We generated six stable individual transgenic lines by over-expressing the potato GWD1 in rice. Transgenic rice grain starch had 9-fold higher 6-phospho (6-P) monoesters and double amounts of 3-phospho (3-P) monoesters, respectively, compared to control grain. The shape and topography of the transgenic starch granules were moderately altered including surface pores and less well defined edges. The gelatinization temperatures of both rice flour and extracted starch were significantly lower than those of the control and hence negatively correlated with the starch phosphate content. The 6-P content was positively correlated with amylose content and relatively long amylopectin chains with DP25-36, and the 3-P content was positively correlated with short chains of DP6-12. The starch pasting temperature, peak viscosity and the breakdown were lower but the setback was higher for transgenic rice flour. The 6-P content was negatively correlated with texture adhesiveness but positively correlated with the cohesiveness of rice flour gels. Our data demonstrate a way forward to employ a starch bioengineering approach for clean modification of starch, opening up completely new applications for rice starch.

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Section: Resultsmentioning

confidence: 99%

“…1) 23, 25 . D-hordein promoters have earlier been demonstrated to direct strong endosperm specific gene expression 47 .…”

Section: Methodsmentioning

confidence: 99%

Highly phosphorylated functionalized rice starch produced by transgenic rice expressing the potato GWD1 gene

Chen

Sun

Zhou

et al. 2017

Sci Rep

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“…Golden Promise) expressing cDNA of the barley hemoglobin gene HvHb1 (accession number: U94968), donated as a plasmid (CA2α) from Dr Robert D. Hill, University of Manitoba (Duff et al ). The cDNA was cloned into either of the vectors pUCE HordD::USER::NOS or pUCE Ubi::USER::NOS by USER cloning (Hebelstrup et al ). Independent transgenic barley lines (UHb) expressing HvHb1 under the maize ubiquitin‐2 promoter were generated by a modified protocol of Agrobacterium ‐mediated transformation, which is described in Carciofi et al ().…”

Section: Methodsmentioning

confidence: 99%

An assessment of the biotechnological use of hemoglobin modulation in cereals

Hebelstrup

Shah

Simpson

et al. 2013

Physiologia Plantarum

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Non-symbiotic hemoglobin (nsHb) genes are ubiquitous in plants, but their biological functions have mostly been studied in model plant species rather than in crops. nsHb influences cell signaling and metabolism by modulating the levels of nitric oxide (NO). Class 1 nsHb is upregulated under hypoxia and is involved in various biotic and abiotic stress responses. Ectopic overexpression of nsHb in Arabidopsis thaliana accelerates development, whilst targeted overexpression in seeds can increase seed yield. Such observations suggest that manipulating nsHb could be a valid biotechnological target. We studied the effects of overexpression of class 1 nsHb in the monocotyledonous crop plant barley (Hordeum vulgare cv. Golden Promise). nsHb was shown to be involved in NO metabolism in barley, as ectopic overexpression reduced the amount of NO released during hypoxia. Further, as in Arabidopsis, nsHb overexpression compromised basal resistance toward pathogens in barley. However, unlike Arabidopsis, nsHb ectopic overexpression delayed growth and development in barley, and seed specific overexpression reduced seed yield. Thus, nsHb overexpression in barley does not seem to be an efficient strategy for increasing yield in cereal crops. These findings highlight the necessity for using actual crop plants rather than laboratory model plants when assessing the effects of biotechnological approaches to crop improvement.

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“…Subsequent treatment of the PCR products with deoxyuridineexcision reagents generates long 3' overhangs designed to complement each other and thereby facilitating their fusion. The combination of this approach with improved cloning techniques has led to the development of an efficient toolbox for transformation in cereals (Hebelstrup et al, 2010).…”

Section: Cloning Strategies and Toolsmentioning

confidence: 99%

The state-of-the-art of grapevine biotechnology and new breeding technologies (NBTS)

et al. 2019

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Context of the review:The manipulation of the genetic basis controlling grapevine adaptation and phenotypic plasticity can be performed either by classical genetics or biotechnologies. In the last 15 years, considerable knowledge has accumulated about the grapevine genome as well as the mechanisms involved in the interaction of the vine with the environment, pests and diseases. Despite the difficulties associated with genetic mapping in this species (allele diversity, chimerism, long generation intervals…), several major QTLs (quantitative trait loci) controlling important vegetative or reproductive traits have been identified. Considering the huge genotypic and phenotypic diversities existing in Vitis, breeding offers a substantial range of options to improve the performances of cultivars. However, even if marker-assisted selection was largely developed to shorten breeding programs, the selection of improved cultivars, whether for agronomic traits or disease tolerances, is still long and uncertain. Moreover, breeding by crossing does not preserve cultivar genetic background, when the wine industry and market are still based on varietal wines. Significance of the review: In grapevine, pioneering biotechnologies were set up in the 1960s to propagate and/or clean the material from micro-organisms. In the 1990s, the basis of genetic engineering was primary established through biolistic or Agrobacterium with several derived technologies refined in the last 10 years. The latest advance is represented by a group of technologies based on genome editing which allows a much more precise modification of the genome. These technologies, so-called NBTs (new breeding technologies), which theoretically do not deconstruct the phenotype of existing cultivars, could be potentially better accepted by the wine industry and consumers than previous GMO (genetically modified organism) approaches. This paper reviews the current state-ofthe-art of the biotechnologies available for grapevine genome manipulation and future prospects for genetic improvement.

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UCE: A uracil excision (USER™)-based toolbox for transformation of cereals

Cited by 22 publications

References 23 publications

Highly phosphorylated functionalized rice starch produced by transgenic rice expressing the potato GWD1 gene

Highly phosphorylated functionalized rice starch produced by transgenic rice expressing the potato GWD1 gene

An assessment of the biotechnological use of hemoglobin modulation in cereals

The state-of-the-art of grapevine biotechnology and new breeding technologies (NBTS)

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