Cold inducible promoter driven Cre- lox system proved to be highly efficient for marker gene excision in transgenic barley

Éva, Csaba; Téglás, Flóra; Zelenyánszki, Helga; Tamás, Cecília; Juhász, Angéla; Mészáros, Katalin; Tamás, László

doi:10.1016/j.jbiotec.2017.10.016

Cited by 19 publications

(11 citation statements)

References 52 publications

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“…Strategies for enzyme delivery vary among studies and are dependent on the goals of each research project. While creative methods for expression have previously been demonstrated, such as use of inducible promoters and/ or autoexcision methods (Éva et al, 2017;Hoff, Schnorr, & Mundy, 2001;Li et al, 2007;Zhang et al, 2003;Zuo, Niu, Moller, & Chua, 2001), most common approaches are transient or stable expression. In the present work, five different recombinase coding sequences were stably integrated into the maize genome through Agrobacterium-mediated transformation.…”

Section: Discussionmentioning

confidence: 99%

Site‐specific recombinase genome engineering toolkit in maize

et al. 2020

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Site‐specific recombinase enzymes function in heterologous cellular environments to initiate strand‐switching reactions between unique DNA sequences termed recombinase binding sites. Depending on binding site position and orientation, reactions result in integrations, excisions, or inversions of targeted DNA sequences in a precise and predictable manner. Here, we established five different stable recombinase expression lines in maize through Agrobacterium‐mediated transformation of T‐DNA molecules that contain coding sequences for Cre, R, FLPe, phiC31 Integrase, and phiC31 excisionase. Through the bombardment of recombinase activated DsRed transient expression constructs, we have determined that all five recombinases are functional in maize plants. These recombinase expression lines could be utilized for a variety of genetic engineering applications, including selectable marker removal, targeted transgene integration into predetermined locations, and gene stacking.

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Section: Discussionmentioning

confidence: 99%

Site‐specific recombinase genome engineering toolkit in maize

et al. 2020

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“…It is also notable, that almost all efforts to improve photosynthetic efficiency involve GMO technology. There is much to done to foster its acceptance [146]. Hopefully biotechnology will come out with reliable solutions for enhancing plant productivity and thereby contributing to solve food and energy crises.…”

Section: Discussionmentioning

confidence: 99%

Current and possible approaches for improving photosynthetic efficiency

2019

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One of the most important tasks laying ahead today's biotechnology is to improve crop productivity with the aim of meeting increased food and energy demands of humankind. Plant productivity depends on many genetic factors, including life cycle, harvest index, stress tolerance and photosynthetic activity. Many approaches were already tested or suggested to improve either. Limitations of photosynthesis have also been uncovered and efforts been taken to increase its efficiency. Examples include decreasing photosynthetic antennae size, 2 increasing the photosynthetically available light spectrum, countering oxygenase activity of Rubisco by implementing C4 photosynthesis to C3 plants and altering source to sink transport of metabolites. A natural and effective photosynthetic adaptation, the sugar alcohol metabolism got however remarkably little attention in the last years, despite being comparably efficient as C4, and can be considered easier to introduce to new species. We also propose root to shoot carbon-dioxide transport as a means to improve photosynthetic performance and drought tolerance at the same time. Different suggestions and successful examples are covered here for improving plant photosynthesis as well as novel perspectives are presented for future research.

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“…The expression of foreign genes in transgenic plants changed when plants were under induction and could affect the excision efficiency in transgenic lines with recombination system. Éva et al obtained transgenic barley with Cre/ loxP system and verified that Cre could accumulate massively in low but be active in higher temperature, low temperature inhibited the premature deletion of the Cre encoding gene, and promote high expression of recombinase therefore increased excision frequency when temperature was high enough to induce Cre enzyme activity [ 54 ]. Our results suggest that a high temperature improves the activation of Gmhsp17.6-L to induce the expression of FLP in order to recognize the loxP-FRT locus and to excise foreign genes between two loxP - FRT sites.…”

Section: Discussionmentioning

confidence: 99%

“…Our results suggest that a high temperature improves the activation of Gmhsp17.6-L to induce the expression of FLP in order to recognize the loxP-FRT locus and to excise foreign genes between two loxP - FRT sites. Incomplete excision occurs possibly because the recombinase activity is affected by gene transcription, temperature, expression levels [ 54 ], or insufficient expression of recombinase, different sensitiveness of plant tissues [ 35 ].…”

Section: Discussionmentioning

confidence: 99%

Heat-Shock-Induced Removal of Transgenes Using the Gene-Deletor System in Hybrid Aspen (Populus tremula × P. tremuloides)

Wang

Zhang

Zhao

et al. 2018

Genes

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To evaluate the efficacy of the gene-deletor system in aspen, we evaluated the system for foreign gene removal in a hybrid aspen clone, INRA 353-53 (Populus tremula × P. tremuloides). The recombinase flipping DNA (FLP) gene was under the control of the heat-inducible promoter of Gmhsp17.6-L, and the β-glucuronidase (gusA) gene which was under the control of the 35S promoter and were constructed using the gene-deletor system in the pCaLFGmFNLFG vector. Six transgenic plants and their sublines were heated at 42 °C for 8 h and gene deletion was verified by polymerase chain reaction (PCR). Three lines exhibited partial transgene deletion while the remaining three lines did not delete. Transgenic lines were evaluated by Southern-blot analyses, verifying that the six transgenic plant lines all had a single copy of transfer DNA (t-DNA). Two partial-deletion lines and two non-deletion lines were analysed for methylation and expression of promoter and recombinase. Hardly any methylation was detected in the Gmhsp17.6-L promoter or recombinase FLP gene sequences, however, the expression of the promoter and recombinase was increased significantly in the partial-deletion compared with the non-deletion line after heat-shock treatment. These results suggest that the excision efficiency had no direct relationship with methylation status of the Gmhsp17.6-L promoter and FLP recombinase, yet was affected by the expression of the Gmhsp17.6-L and FLP after heat-shock treatment.

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Cold inducible promoter driven Cre- lox system proved to be highly efficient for marker gene excision in transgenic barley

Cited by 19 publications

References 52 publications

Site‐specific recombinase genome engineering toolkit in maize

Site‐specific recombinase genome engineering toolkit in maize

Current and possible approaches for improving photosynthetic efficiency

Heat-Shock-Induced Removal of Transgenes Using the Gene-Deletor System in Hybrid Aspen (Populus tremula × P. tremuloides)

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