Fast, efficient and reproducible genetic transformation of Phaseolus spp. by Agrobacterium rhizogenes

Estrada-Navarrete, Georgina; Alvarado‐Affantranger, Xóchitl; Olivares, Juan-Elías; Guillén, Gabriel; Díaz-Camino, Claudia; Campos, Francisco; Quinto, Carmen; Gresshoff, Peter M.; Sánchez, Federico

doi:10.1038/nprot.2007.259

Cited by 112 publications

(118 citation statements)

References 15 publications

Supporting

Mentioning

114

Contrasting

Unclassified

Order By: Relevance

“…This protocol has been used as an alternative method for stable transformation in common bean and other recalcitrant species (Estrada-Navarrete et al, 2007). The construct for miR172 overexpression (OE172) contained the 35S cauliflower mosaic virus promoter fused to the miR172c precursor.…”

Section: Effect Of Mir172c Overexpression On Root Development and Rhimentioning

confidence: 99%

See 1 more Smart Citation

The Micro-RNA172c-APETALA2-1 Node as a Key Regulator of the Common Bean-Rhizobium etli Nitrogen Fixation Symbiosis

et al. 2015

Self Cite

View full text Add to dashboard Cite

Micro-RNAs are recognized as important posttranscriptional regulators in plants. The relevance of micro-RNAs as regulators of the legume-rhizobia nitrogen-fixing symbiosis is emerging. The objective of this work was to functionally characterize the role of micro-RNA172 (miR172) and its conserved target APETALA2 (AP2) transcription factor in the common bean (Phaseolus vulgaris)-Rhizobium etli symbiosis. Our expression analysis revealed that mature miR172c increased upon rhizobial infection and continued increasing during nodule development, reaching its maximum in mature nodules and decaying in senescent nodules. The expression of AP2-1 target showed a negative correlation with miR172c expression. A drastic decrease in miR172c and high AP2-1 mRNA levels were observed in ineffective nodules. Phenotypic analysis of composite bean plants with transgenic roots overexpressing miR172c or a mutated AP2-1 insensitive to miR172c cleavage demonstrated the pivotal regulatory role of the miR172 node in the common beanrhizobia symbiosis. Increased miR172 resulted in improved root growth, increased rhizobial infection, increased expression of early nodulation and autoregulation of nodulation genes, and improved nodulation and nitrogen fixation. In addition, these plants showed decreased sensitivity to nitrate inhibition of nodulation. Through transcriptome analysis, we identified 114 common bean genes that coexpressed with AP2-1 and proposed these as being targets for transcriptional activation by AP2-1. Several of these genes are related to nodule senescence, and we propose that they have to be silenced, through miR172c-induced AP2-1 cleavage, in active mature nodules. Our work sets the basis for exploring the miR172-mediated improvement of symbiotic nitrogen fixation in common bean, the most important grain legume for human consumption.

show abstract

Section: Effect Of Mir172c Overexpression On Root Development and Rhimentioning

confidence: 99%

“…Common bean composite plants with transformed root system and untransformed aerial system were generated as described (Estrada-Navarrete et al, 2007;Aparicio-Fabre et al, 2013). For plant transformation, Agrobacterium rhizogenes K599 strains bearing the EV, OE172, or OEAP2m plasmids were used.…”

Section: Plasmid Construction Plant Transformation and Production Omentioning

confidence: 99%

The Micro-RNA172c-APETALA2-1 Node as a Key Regulator of the Common Bean-Rhizobium etli Nitrogen Fixation Symbiosis

et al. 2015

Self Cite

View full text Add to dashboard Cite

show abstract

“…Common bean seeds (Phaseolus vulgaris cv Negro Jamapa) were used for Agrobacterium rhizogenes K599-mediated transformation to generate hairy roots in composite plants using a previously described method (Estrada-Navarrete et al, 2007). Usually at the second week after inoculation, hairy roots emerging from A. rhizogenes K599-induced calli are ;3 cm long.…”

Section: Bean Hairy Root Transformationmentioning

confidence: 99%

“…To evaluate the spatiotemporal expression of Pv-PI3K during nodule organogenesis, we cloned the Pv-PI3K promoter (region ;1 kb directly upstream of the initiation codon) in a transcriptional fusion with the chimeric reporter GFP-GUS to yield PvPI3K pro :GFP-GUS. Pv-PI3K promoter activity was therefore monitored as GUS activity or GFP fluorescence in P. vulgaris transgenic hairy roots generated by Agrobacterium rhizogenes K599-mediated transformation (Estrada-Navarrete et al, 2007). Control transgenic roots harbored a cassette with no promoter sequences upstream of the GFP-GUS sequences.…”

Section: Pv-pi3k Transcript Levels During the Symbiosis Between P Vumentioning

confidence: 99%

An Autophagy-Related Kinase Is Essential for the Symbiotic Relationship between Phaseolus vulgaris and Both Rhizobia and Arbuscular Mycorrhizal Fungi

Estrada-Navarrete

Cruz-Mireles

Lascano³

et al. 2016

Plant Cell

Self Cite

View full text Add to dashboard Cite

Eukaryotes contain three types of lipid kinases that belong to the phosphatidylinositol 3-kinase (PI3K) family. In plants and Saccharomyces cerevisiae, only PI3K class III family members have been identified. These enzymes regulate the innate immune response, intracellular trafficking, autophagy, and senescence. Here, we report that RNAi-mediated downregulation of common bean (Phaseolus vulgaris) PI3K severely impaired symbiosis in composite P. vulgaris plants with endosymbionts such as Rhizobium tropici and Rhizophagus irregularis. Downregulation of Pv-PI3K was associated with a marked decrease in root hair growth and curling. Additionally, infection thread growth, root-nodule number, and symbiosome formation in root nodule cells were severely affected. Interestingly, root colonization by AM fungi and the formation of arbuscules were also abolished in PI3K loss-of-function plants. Furthermore, the transcript accumulation of genes encoding proteins known to interact with PI3K to form protein complexes involved in autophagy was drastically reduced in these transgenic roots. RNAimediated downregulation of one of these genes, Beclin1/Atg6, resulted in a similar phenotype as observed for transgenic roots in which Pv-PI3K had been downregulated. Our findings show that an autophagy-related process is crucial for the mutualistic interactions of P. vulgaris with beneficial microorganisms.

show abstract

“…The Agrobacterium rhizogenes K599 strain carrying the corresponding constructs was used to generate hairy root formation on bean tissues and to form composite plants after transformation (Estrada-Navarrete et al, 2007). Transgenic hairy roots expressing the TOR-RNAi vectors and the TOR::GFP-GUS promoter were selected using an epifluorescence stereomicroscope based on the fluorescence of red fluorescent protein (RFP) and GFP, respectively.…”

Section: Plasmid Construction and Composite Plantsmentioning

confidence: 99%

A Legume TOR Protein Kinase Regulates Rhizobium Symbiosis and Is Essential for Infection and Nodule Development

et al. 2016

Self Cite

View full text Add to dashboard Cite

The target of rapamycin (TOR) protein kinase regulates metabolism, growth, and life span in yeast, animals, and plants in coordination with nutrient status and environmental conditions. The nutrient-dependent nature of TOR functionality makes this kinase a putative regulator of symbiotic associations involving nutrient acquisition. However, TOR's role in these processes remains to be understood. Here, we uncovered the role of TOR during the bean (Phaseolus vulgaris)-Rhizobium tropici (Rhizobium) symbiotic interaction. TOR was expressed in all tested bean tissues, with higher transcript levels in the root meristems and senesced nodules. We showed TOR promoter expression along the progressing infection thread and in the infected cells of mature nodules. Posttranscriptional gene silencing of TOR using RNA interference (RNAi) showed that this gene is involved in lateral root elongation and root cell organization and also alters the density, size, and number of root hairs. The suppression of TOR transcripts also affected infection thread progression and associated cortical cell divisions, resulting in a drastic reduction of nodule numbers. TOR-RNAi resulted in reduced reactive oxygen species accumulation and altered CyclinD1 and CyclinD3 expression, which are crucial factors for infection thread progression and nodule organogenesis. Enhanced expression of TORregulated ATG genes in TOR-RNAi roots suggested that TOR plays a role in the recognition of Rhizobium as a symbiont. Together, these data suggest that TOR plays a vital role in the establishment of root nodule symbiosis in the common bean.In plants, growth occurs as a result of cell growth coupled with cell proliferation and cell expansion caused by nutrient availability. Unlike animals, in plants, postembryonic growth is sustained by specialized zones (shoot meristems and root apices) and depends upon the availability of nutrients that are present externally or generated by various cellular processes. These local and systemic nutrient and energy sources are integrated, and information is communicated to metabolic regulators through signaling networks to control cellular processes.The evolutionarily conserved target of rapamycin (TOR) is a Ser/Thr kinase that acts as a master regulator of metabolism and growth. In mammals, TOR exerts translational control of cell proliferation (Dowling et al., 2010), tumorigenesis, metastasis (Hsieh et al., 2012), and insulin signaling (Hsu et al., 2011). Loss-of-function TOR mutations in Saccharomyces cerevisiae and Schizosaccharomyces pombe (Barbet et al., 1996;Weisman and Choder, 2001), Drosophila melanogaster (Zhang et al., 2000), Caenorhabditis elegans (Long et al., 2002), and mice (Murakami et al., 2004) resulted in embryo lethality and other growth-associated problems.Unlike other eukaryotes, knowledge about TOR in plants is limited due to a lack of molecular and biochemical assays for endogenous TOR kinase activity.

show abstract

Fast, efficient and reproducible genetic transformation of Phaseolus spp. by Agrobacterium rhizogenes

Cited by 112 publications

References 15 publications

The Micro-RNA172c-APETALA2-1 Node as a Key Regulator of the Common Bean-Rhizobium etli Nitrogen Fixation Symbiosis

The Micro-RNA172c-APETALA2-1 Node as a Key Regulator of the Common Bean-Rhizobium etli Nitrogen Fixation Symbiosis

An Autophagy-Related Kinase Is Essential for the Symbiotic Relationship between Phaseolus vulgaris and Both Rhizobia and Arbuscular Mycorrhizal Fungi

A Legume TOR Protein Kinase Regulates Rhizobium Symbiosis and Is Essential for Infection and Nodule Development

Contact Info

Product

Resources

About