Agrobacterium rhizogenes—mediated transformation ofPisum sativumL. roots as a tool for studying the mycorrhizal and root nodule symbioses

Abstract

In this study, we demonstrated the successful transformation of two pea (Pisum sativum L.) cultivars using Agrobacterium rhizogenes, whereby transgenic roots in the resulting composite plants showed expression of the gene encoding the green fluorescent protein. Subsequent to infection with A. rhizogenes, approximately 70%–80% of pea seedlings developed transgenic hairy roots. We found out that the transgenic roots can be efficiently nodulated by Rhizobium leguminosarum bv. viciae and infected by the arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis. The morphology of nodules in the transgenic roots was found to be identical to that of nodules observed in wild-type roots, and we also observed the effective induction of markers typical of the symbiotic association with AM fungi. The convenient protocol for highly efficient A. rhizogenes-mediated transformation developed in this study would be a rapid and effective tool for investigating those genes involved in the development of the two types of symbioses found in pea plants.

High-Efficiency Agrobacterium rhizogenes-Mediated Transgenic Hairy Root Induction of Lens culinaris

“…As an alternative, the A. rhizogenes-mediated generation of composite plants, consisting of transgenic roots and wild-type shoot tissues, has proven to be an amenable approach to rapidly generate hairy roots carrying transgenes of interest, as evidenced in a wide range of plant species. In legumes, A. rhizogenes-mediated transformation was first researched in Lotus corniculantus to facilitate studies on nodule formation [17,36], while later efforts focused on the development of related protocols in model plants [22,23] as well as cultivated species [25,28,32,33]. Given the lack of relative transformation protocols in lentil, this study aimed at optimizing a protocol enabling an efficient generation of transgenic roots to serve as a suitable platform for root biology studies.…”

“…In legumes, the production of Ri T-DNA-transformed hairy roots has been realized with a variety of bacterial strains. Strain K599 has proven capable of transforming Phaseolus vulgaris [33], Glycine max [37] and Cicer arietinum [32], while strain Arqua has successfully transformed Medicago truncatula [38] and Pisium sativum [28]. Both Arqua and K599 are considered to be A. rhizogenes strains of low virulence, thus eliciting a limited number of hairy roots which exhibit a growth pattern comparable to wild-type roots [19,39].…”

Studying the effect of tissue-specific expression of theK1gene encoding LysM-receptor-like kinase on the development of symbiosis in peas

“…The transformation of plants was performed using the method described earlier [21]. Histochemical staining of roots was performed using the method described earlier [12].…”

Effect of the co-inoculation of Burkholderia ubonensis and Rhizobium spp. in the nodulation of Pisum sativum

“…Esta leguminosa, cultivada en todo el mundo, aporta significativamente a la agricultura sostenible al tener un ciclo de vida corto y requerir bajas cantidades de agua para su cultivo; además, es muy valorado su aporte de nitrógeno orgánico al suelo a través de la fijación biológica del nitrógeno en asociaciones con bacterias (Do Carmo et al, 2016;Xiong et al, 2018;Pietrysiak et al, 2018). Las leguminosas están involucradas en asociaciones simbióticas con un grupo heterogéneo de bacterias rizobianas (Leppyanen et al, 2019), ubicadas dentro de las proteobacterias α y β; siendo su principal fuente de aislamiento los nódulos radiculares. Estas bacterias tienen un rango de hospedero muy estrecho y forman nódulos con plantas bien definidas; son diversos los estudios que confirman que el simbionte de P. sativum es Rhizobium leguminosarum bv.…”

“…Estas bacterias tienen un rango de hospedero muy estrecho y forman nódulos con plantas bien definidas; son diversos los estudios que confirman que el simbionte de P. sativum es Rhizobium leguminosarum bv. Viciae, un bacilo aeróbico, móvil, no formador de esporas y perteneciente al grupo de los α-rizobios (Kumar et al, 2015;Leppyanen et al, 2019;Bourion et al, 2018;Mazur et al, 2015;Reeve et al, 2015).…”