Application of the acetolactate synthase gene as a cisgenic selectable marker for Agrobacterium-mediated transformation in Chinese cabbage (Brassica rapa ssp. pekinensis)

Konagaya, Ken Ichi; Tsuda, Masayuki; Okuzaki, Ayako; Ando, Sugihiro; Tabei, Yutaka

doi:10.5511/plantbiotechnology.13.0124a

Cited by 11 publications

(12 citation statements)

References 42 publications

(49 reference statements)

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“…The generation of cisgenic crops is still very limited and has been reported in apple (Vanblaere et al, 2011 , 2014 ), in barley (Holme et al, 2012 ), and in potato (Jo et al, 2014 ). In many crops the first steps toward the production of cisgenic plants are reported (Corredoira et al, 2012 ; An et al, 2013 ; Konagaya et al, 2013 ), e.g., in Rosaceae tree species such as pear (Righetti et al, 2014 ) and apple (Joshi et al, 2011 ; Würdig et al, 2013 ). In the latter crop, the first steps toward stacking multiple cisgenes and herewith multiple resistances have been taken (Gessler et al, 2014 ).…”

Section: Introductionmentioning

confidence: 99%

Cisgenic apple trees; development, characterization, and performance

et al. 2015

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Two methods were developed for the generation of cisgenic apples. Both have been successfully applied producing trees. The first method avoids the use of any foreign selectable marker genes; only the gene-of-interest is integrated between the T-DNA border sequences. The second method makes use of recombinase-based marker excision. For the first method we used the MdMYB10 gene from a red-fleshed apple coding for a transcription factor involved in regulating anthocyanin biosynthesis. Red plantlets were obtained and presence of the cisgene was confirmed. Plantlets were grafted and grown in a greenhouse. After 3 years, the first flowers appeared, showing red petals. Pollination led to production of red-fleshed cisgenic apples. The second method used the pM(arker)F(ree) vector system, introducing the scab resistance gene Rvi6, derived from apple. Agrobacterium-mediated transformation, followed by selection on kanamycin, produced genetically modified apple lines. Next, leaves from in vitro material were treated to activate the recombinase leading to excision of selection genes. Subsequently, the leaf explants were subjected to negative selection for marker-free plantlets by inducing regeneration on medium containing 5-fluorocytosine. After verification of the marker-free nature, the obtained plants were grafted onto rootstocks. Young trees from four cisgenic lines and one intragenic line, all containing Rvi6, were planted in an orchard. Appropriate controls were incorporated in this trial. We scored scab incidence for three consecutive years on leaves after inoculations with Rvi6-avirulent strains. One cisgenic line and the intragenic line performed as well as the resistant control. In 2014 trees started to overcome their juvenile character and formed flowers and fruits. The first results of scoring scab symptoms on apple fruits were obtained. Apple fruits from susceptible controls showed scab symptoms, while fruits from cisgenic and intragenic lines were free of scab.

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Section: Introductionmentioning

confidence: 99%

Cisgenic apple trees; development, characterization, and performance

et al. 2015

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“…Kanamycin was used in this experiment as a selective agent. But in other plant species, for example Pisum sativum, Cucumis sativus, Jatropha curcus and Brassica rapa, some herbicides such as glyphosate, bialaphos, and bispyribac are more effective for producing transgenic plants (Bean et al 1997;Kajikawa et al 2012;Konagaya et al 2013;Vengadesan et al 2005). An efficient transformation system for other Cucurbita species may be established by optimization of the factors described above.…”

Section: Conclusion and Future Perspectives For Transformation Of Cumentioning

confidence: 99%

Improving the transformation efficiency of Cucurbita species: factors and strategy for practical application

Nanasato

Okuzaki

Tabei

2013

Plant Biotechnology

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Cucurbita species are refractory to transformation. Hence, the only 2 reports published regarding the transformation of Cucurbita species until our successful transformation of C. moschata (cv. Heiankogiku) in 2011. The efficiency was 2.7±1.3% using wounded explants vortexed with whisker suspension. To improve transformation efficiency, transformation experiments were carried out with various ages of cotyledonary explants. The highest efficiency was obtained with 1-day-old (3.2±0.9%) and 2-day-old (3.3±0.8%) explants after germination. Histochemical analysis of GUS activity revealed that wounding allowed Agrobacterium access to the deeper layer of explants. These results suggested that cells having a high ability of shoot organogenesis exist not on the surface but in the deeper layers of explants. We applied vacuum infiltration to wounded explants to enhance Agrobacterium access to the deeper layers, which resulted in improvement of transformation efficiency by 3-fold (9.2±2.9%). To verify the efficacy of our transformation procedure for other Cucurbita species, we attempted to obtain transformants with 3 Cucurbita species: C. maxima (cv. Ebisu), C. pepo (cv. Black Tosca), and C. ficifolia (cv. Kurotanenankin). The average regeneration efficiencies were 54.2±7.2%, 62.5±19.1%, and 72.2±13.4% under our regeneration system. Although the efficiency for production of tansgenic plants was very low (ca. 0.2-0.3%), a transgenic line was obtained from C. maxima and C. pepo, respectively. We discuss recent advances that may help in the development of beneficial applications for the transformation of Cucurbita species.

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“…Resistance to acetolactate synthase (ALS) inhibiting herbicides was established via ODM in oilseed rape (Gocal et al, 2015 ), by SDN-1 technology in potato with TALEN (Nicolia et al, 2015 ; Butler et al, 2016 ) and CRISPR/Cas9 (Butler et al, 2016 ), in rice with TALEN (Li et al, 2016a ) and in tobacco with ZNF (Townsend et al, 2009 ). In Chinese cabbage this trait was introduced by cisgenesis (Konagaya et al, 2013 ).…”

Section: Risk Assessment Considerationsmentioning

confidence: 99%

An EU Perspective on Biosafety Considerations for Plants Developed by Genome Editing and Other New Genetic Modification Techniques (nGMs)

Eckerstorfer

Dolezel

Heissenberger

et al. 2019

Front. Bioeng. Biotechnol.

101

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The question whether new genetic modification techniques (nGM) in plant development might result in non-negligible negative effects for the environment and/or health is significant for the discussion concerning their regulation. However, current knowledge to address this issue is limited for most nGMs, particularly for recently developed nGMs, like genome editing, and their newly emerging variations, e.g., base editing. This leads to uncertainties regarding the risk/safety-status of plants which are developed with a broad range of different nGMs, especially genome editing, and other nGMs such as cisgenesis, transgrafting, haploid induction or reverse breeding. A literature survey was conducted to identify plants developed by nGMs which are relevant for future agricultural use. Such nGM plants were analyzed for hazards associated either (i) with their developed traits and their use or (ii) with unintended changes resulting from the nGMs or other methods applied during breeding. Several traits are likely to become particularly relevant in the future for nGM plants, namely herbicide resistance (HR), resistance to different plant pathogens as well as modified composition, morphology, fitness (e.g., increased resistance to cold/frost, drought, or salinity) or modified reproductive characteristics. Some traits such as resistance to certain herbicides are already known from existing GM crops and their previous assessments identified issues of concern and/or risks, such as the development of herbicide resistant weeds. Other traits in nGM plants are novel; meaning they are not present in agricultural plants currently cultivated with a history of safe use, and their underlying physiological mechanisms are not yet sufficiently elucidated. Characteristics of some genome editing applications, e.g., the small extent of genomic sequence change and their higher targeting efficiency, i.e., precision, cannot be considered an indication of safety per se, especially in relation to novel traits created by such modifications. All nGMs considered here can result in unintended changes of different types and frequencies. However, the rapid development of nGM plants can compromise the detection and elimination of unintended effects. Thus, a case-specific premarket risk assessment should be conducted for nGM plants, including an appropriate molecular characterization to identify unintended changes and/or confirm the absence of unwanted transgenic sequences.

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Application of the acetolactate synthase gene as a cisgenic selectable marker for Agrobacterium-mediated transformation in Chinese cabbage (Brassica rapa ssp. pekinensis)

Cited by 11 publications

References 42 publications

Cisgenic apple trees; development, characterization, and performance

Cisgenic apple trees; development, characterization, and performance

Improving the transformation efficiency of Cucurbita species: factors and strategy for practical application

An EU Perspective on Biosafety Considerations for Plants Developed by Genome Editing and Other New Genetic Modification Techniques (nGMs)

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