Non-antibiotic selection systems for soybean somatic embryos: the lysine analog aminoethyl-cysteine as a selection agent

Rao, Suguna; Mamadou, Lewamy; McConnell, Matthew; Polisetty, Raghuveer; Kwanyuen, P.; Hildebrand, David F.

doi:10.1186/1472-6750-9-94

Cited by 5 publications

(3 citation statements)

References 58 publications

(68 reference statements)

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“…A similar low concentration of chlorsulfuron (10 nM) has previously been found to effectively inhibit transgenic rice protoplast growth (Li et al 1992). On the other hand, chlorsulfuron-resistant maize calli were selected on 50 nM chlorsulfuron, while soybean embryos were only sensitive to ALS herbicides (mix of primisulfuron and prosulfuron) when a concentration as high as 183 mg/l was applied (Fromm et al 1990; Rao et al 2009). …”

Section: Discussionmentioning

confidence: 99%

In vitro selection of transgenic sugarcane callus utilizing a plant gene encoding a mutant form of acetolactate synthase

Vyver

Conradie

Koßmann

et al. 2013

In Vitro Cell.Dev.Biol.-Plant

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Selection genes are routinely used in plant genetic transformation protocols to ensure the survival of transformed cells by limiting the regeneration of non-transgenic cells. In order to find alternatives to the use of antibiotics as selection agents, we followed a targeted approach utilizing a plant gene, encoding a mutant form of the enzyme acetolactate synthase, to convey resistance to herbicides. The sensitivity of sugarcane callus (Saccharum spp. hybrids, cv. NCo310) to a number of herbicides from the sulfonylurea and imidazolinone classes was tested. Callus growth was most affected by sulfonylurea herbicides, particularly 3.6 μg/l chlorsulfuron. Herbicide-resistant transgenic sugarcane plants containing mutant forms of a tobacco acetolactate synthase (als) gene were obtained following biolistic transformation. Post-bombardment, putative transgenic callus was selectively proliferated on MS medium containing 3 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D), 20 g/l sucrose, 0.5 g/l casein, and 3.6 μg/l chlorsulfuron. Plant regeneration and rooting was done on MS medium lacking 2,4-D under similar selection conditions. Thirty vigorously growing putative transgenic plants were successfully ex vitro-acclimatized and established under glasshouse conditions. Glasshouse spraying of putative transgenic plants with 100 mg/l chlorsulfuron dramatically decreased the amount of non-transgenic plants that had escaped the in vitro selection regime. PCR analysis showed that six surviving plants were als-positive and that five of these expressed the mutant als gene. This report is the first to describe a selection system for sugarcane transformation that uses a selectable marker gene of plant origin targeted by a sulfonylurea herbicide.

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Section: Discussionmentioning

confidence: 99%

In vitro selection of transgenic sugarcane callus utilizing a plant gene encoding a mutant form of acetolactate synthase

Vyver

Conradie

Koßmann

et al. 2013

In Vitro Cell.Dev.Biol.-Plant

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“…Paz et al (2006) used the cotyledonary node of half-seeds as an explant and reported 1.4-8.7% transformation efficiency. Recently, Rao et al (2009) reported the selection of soybean somatic embryos on non-antibiotic selection medium as lysine-insensitive dihydrodipicolinate synthase gene with non-antibiotic selection agent S-(2-aminoethyl)-Lcysteine.…”

Section: Introductionmentioning

confidence: 99%

Expression of rol genes in transgenic soybean (Glycine max L.) leads to changes in plant phenotype, leaf morphology, and flowering time

Zia

Mirza

Malik

et al. 2010

Plant Cell Tiss Organ Cult

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Soybean (Glycine max L.) cultivar NARC-4 was transformed with constructs carrying rolA, rolB, or rolC genes, each under the control of the Cauliflower Mosaic Virus 70S promoter. Cotyledonary nodes of soybean seeds were infected with Agrobacterium tumefaciens strain LBA4404 carrying one of the three rol genes, along with nptII in the plasmid pLBR. The efficiency of the transformation varied slightly among the three constructs, with frequencies of 6, 7, and 5% for the rolA, rolB, and rolC genes, respectively, being observed. Southern blot analysis confirmed the integration of rol genes in the soybean genome with varying numbers of copies of the transgene. All transformed plants showed enhanced rooting, but the number of adventitious roots was higher for transformants carrying the rolB transgene. rolA and rolC transformants showed dwarf phenotypes, clustered branching, and variations in leaf morphology. Furthermore, these plants flowered within a short period of time and produced lower numbers of flowers. rolB transformants showed variations in phenotype, including dwarf to semidwarf and shrubby growth, abnormal stem growth, short internodes, variations in leaf morphology, and greenish to yellowish-green colored leaves. These plants also flowered early, but dwarf plants produced low numbers of flowers, while shrubby plants produced high numbers of flowers, but these were mostly infertile.

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“…Preliminary results in our lab showed that this marker/agent can also efficiently select transgenic somatic embryos (unpublished results). Rao et al . (2009) successfully developed a system to select somatic embryos using the E. coli dap A gene, which confers resistance to glufosinate, glyphosate, S-(2 aminoethyl)-L-cysteine and imidazolinones.…”

Section: Selection Systemmentioning

confidence: 99%

Soybean genetic transformation: a valuable tool for the functional study of genes and the production of agronomically improved plants

et al. 2012

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Transgenic plants represent an invaluable tool for molecular, genetic, biochemical and physiological studies by gene overexpression or silencing, transposon-based mutagenesis, protein sub-cellular localization and/or promoter characterization as well as a breakthrough for breeding programs, allowing the production of novel and genetically diverse genotypes. However, the stable transformation of soybean cannot yet be considered to be routine because it depends on the ability to combine efficient transformation and regeneration techniques. Two methods have been used with relative success to produce completely and stably transformed plants: particle bombardment and the Agrobacterium tumefaciens system. In addition, transformation by Agrobacterium rhizogenes has been used as a powerful tool for functional studies. Most available information on gene function is based on heterologous expression systems. However, as the activity of many promoters or proteins frequently depends on specific interactions that only occur in homologous backgrounds, a final confirmation based on a homologous expression system is desirable. With respect to soybean biotech improvement, transgenic lines with agronomical, nutritional and pharmaceutical traits have been obtained, including herbicide-tolerant soybeans, which represented the principal biotech crop in 2011, occupying 47% of the global biotech area.

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Non-antibiotic selection systems for soybean somatic embryos: the lysine analog aminoethyl-cysteine as a selection agent

Cited by 5 publications

References 58 publications

In vitro selection of transgenic sugarcane callus utilizing a plant gene encoding a mutant form of acetolactate synthase

In vitro selection of transgenic sugarcane callus utilizing a plant gene encoding a mutant form of acetolactate synthase

Expression of rol genes in transgenic soybean (Glycine max L.) leads to changes in plant phenotype, leaf morphology, and flowering time

Soybean genetic transformation: a valuable tool for the functional study of genes and the production of agronomically improved plants

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