Molecular cloning and characterization of 5-enolpyruvylshikimate-3-phosphate synthase gene from Convolvulus arvensis L.

Huang, Zhengxu; Zhang, Chaoxian; Huang, Hongjuan; Wei, Shugang; Liu, Yan; Cui, Hailan; Chen, Jingchao; Yang, Long; Chen, Jinyi

doi:10.1007/s11033-014-3056-2

Cited by 10 publications

(12 citation statements)

References 25 publications

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“…The transcription level showed that the aroA gene is expressed extensively in roots, stems, and leaves, being most highly expressed in the leaf. This expression model is consistent with the aroA expression observed in Convolvulus arvensis L. and Camptotheca acuminate (Gong et al, 2006;Huang et al, 2014). Glyphosate treatment could induce a significant increase of aroA gene expression level.…”

Section: Discussionsupporting

confidence: 89%

“…In this experiment, the peak induction was 36 h after spraying glyphosate, which was 3-fold higher than the control group's expression level. In field bindweed, the expression level of EPSPS was also responsive to glyphosate (Huang et al, 2014). These results show that the aroA gene plays an important role in the response to glyphosate.…”

Section: Discussionmentioning

confidence: 62%

“…EPSPS proteins have been divided into two major types: class I and class II (He et al, 2001). Class I EPSPS is found in all plants and microorganisms, including Convolvulus arvensis, Escherichia coli, Proteus mirabilis, and Klebsiella pneumoniae (Rogers et al, 1983;Sost et al, 1990;Wang et al, 2003;Funke et al, 2009;Huang et al, 2014;Tian et al, 2014) and is sensitive to glyphosate. Class II enzymes are found in Pseudomonas stutzeri A1501, Agrobacterium tumefaciens strain CP4, Halothermothrix orenii H168, and Bacillus cereus (Li et al, 2009;Tian et al, 2012Tian et al, , 2013.…”

Section: Introductionmentioning

confidence: 99%

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Expression of a bacterial aroA gene confers tolerance to glyphosate in tobacco plants

Liu

Cao

2018

Turk J Biol

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Glyphosate is a widely used herbicide that inhibits the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS)-encoding aroA gene in the shikimate pathway. The discovery and cloning of the aroA gene with high resistance is central to breeding a transgenic glyphosate-resistant plant. A novel aroA Pantoea gene from Pantoea G-1 was previously isolated and cloned. The aroA Pantoea enzyme was defined as a new class I EPSPS with glyphosate resistance. The aroA Pantoea gene was introduced into tobacco through Agrobacteriummediated transformation. The transgenic tobacco plants were confirmed by PCR, RT-PCR, and Southern blot. The analysis of glyphosate resistance also showed that the transgenic tobacco plants could survive at 15 mM glyphosate; the glyphosate resistance level of the transgenic plants is higher than the agricultural application level recommended by most manufacturers. Overall, this study shows that aroA Pantoea can be used as a candidate gene for the development of genetically modified crops.

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Section: Discussionsupporting

confidence: 89%

Section: Discussionmentioning

confidence: 62%

Section: Introductionmentioning

confidence: 99%

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Expression of a bacterial aroA gene confers tolerance to glyphosate in tobacco plants

Liu

Cao

2018

Turk J Biol

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“…Several plants with different levels of natural glyphosate tolerance have been identified (Boerboom et al 1990;Culpepper 2006;DeGennaro and Weller 1984;Huang et al 2014;Owen and Zelaya 2005;Westhoven et al 2008;Yuan et al 2001). Among them, D. chinensis survived glyphosate at 3.3 kg ha -1 .…”

Section: Discussionmentioning

confidence: 99%

Multiple mechanism confers natural tolerance of three lilyturf species to glyphosate

Mao

Xie

Chen

et al. 2015

Planta

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A combination of unique EPSPS structure and increased gene copy number and expression contribute to natural glyphosate tolerance in three lilyturf species. A few plants are naturally tolerant to glyphosate, the most widely used non-selective herbicide worldwide. Here, the basis for natural tolerance to glyphosate in three lilyturf species, Ophiopogon japonicus (OJ), Liriope spicata (LS), and Liriope platyphylla (LP), is characterized. These species tolerate glyphosate at about five times the commercially recommended field dose. They share three unique amino acids in their 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) that affect glyphosate binding. These correspond to Asp71Met, Ala112Ile, and Val201Met amino acid variations compared to 231 other published plant EPSPS amino acid sequences. There was also a common deletion at 91 of a highly conserved glutamic acid. Glyphosate-treated lilyturf plants accumulated little shikimic acid but had significantly higher levels of EPSPS mRNA than initially expressed in the control. The IC50 of LsEPSPS was 14.0 µM compared to the 5.1 µM of Arabidopsis thaliana. The higher K m and K i values of LsEPSPS kinetics showed that LsEPSPS had lower substrate binding affinity to glyphosate. Overexpression of LsEPSPS in the recombinant E. coli BL21 (DE3) strain enhanced its tolerance to glyphosate. Both OJ and LS had two copies of the EPSPS gene, while LP had three copies. Therefore, a combination of unique EPSPS structure and increased gene copy number and expression contribute to natural glyphosate tolerance in the three lilyturf species.

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“…More recently, it has been shown that Arabidopsis plants transformed with an Arabidopsis EPSPS gene driven by the CaMV35S promoter was tolerant to glyphosate [ 37 ]. In another study, it was shown that a an EPSPS gene from Convolvulus arvensis driven by the CaMV35S promoter also conferred resistance to glyphosate in Arabidopsis [ 38 ]. The Arabidopsis transformants in either study, were not checked for EPSPS protein levels relative to the control plants thus making it difficult to compare the outcome of the present study with those previous studies [ 37 , 38 ].…”

Section: Discussionmentioning

confidence: 99%

An intragenic approach to confer glyphosate resistance in chile (Capsicum annuum) by introducing an in vitro mutagenized chile EPSPS gene encoding for a glyphosate resistant EPSPS protein

et al. 2018

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Chile pepper (Capsicum annuum) is an important high valued crop worldwide, and when grown on a large scale has problems with weeds. One important herbicide used is glyphosate. Glyphosate inactivates the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), a key enzyme in the synthesis of aromatic amino acids. A transgenic approach towards making glyphosate resistant plants, entails introducing copies of a gene encoding for glyphosate-resistant EPSPS enzyme into the plant. The main objective of our work was to use an intragenic approach to confer resistance to glyphosate in chile which would require using only chile genes for transformation including the selectable marker. Tobacco was used as the transgenic system to identify different gene constructs that would allow for the development of the intragenic system for chile, since chile transformation is inefficient. An EPSPS gene was isolated from chile and mutagenized to introduce substitutions that are known to make the encoded enzyme resistant to glyphosate. The promoter for EPSPS gene was isolated from chile and the mutagenized chile EPSPS cDNA was engineered behind both the CaMV35S promoter and the EPSPS promoter. The leaves from the transformants were checked for resistance to glyphosate using a cut leaf assay. In tobacco, though both gene constructs exhibited some degree of resistance to glyphosate, the construct with the CaMV35S promoter was more effective and as such chile was transformed with this gene construct. The chile transformants showed resistance to low concentrations of glyphosate. Furthermore, preliminary studies showed that the mutated EPSPS gene driven by the CaMV35S promoter could be used as a selectable marker for transformation. We have shown that an intragenic approach can be used to confer glyphosate-resistance in chile. However, we need a stronger chile promoter and a mutated chile gene that encodes for a more glyphosate resistant EPSPS protein.

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Molecular cloning and characterization of 5-enolpyruvylshikimate-3-phosphate synthase gene from Convolvulus arvensis L.

Cited by 10 publications

References 25 publications

Expression of a bacterial aroA gene confers tolerance to glyphosate in tobacco plants

Expression of a bacterial aroA gene confers tolerance to glyphosate in tobacco plants

Multiple mechanism confers natural tolerance of three lilyturf species to glyphosate

An intragenic approach to confer glyphosate resistance in chile (Capsicum annuum) by introducing an in vitro mutagenized chile EPSPS gene encoding for a glyphosate resistant EPSPS protein

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