Use of Agrobacterium expressing green fluorescent protein to evaluate colonization of sonication-assisted Agrobacterium-mediated transformation-treated soybean cotyledons

Abstract

Colonization and infection of soybean cotyledons by Agrobacterium tumefaciens and subsequent elimination of bacteria from cotyledons were monitored using bacteria expressing green fluorescent protein (GFP). GFP provided a quick, non‐destructive method to evaluate, in real time, Agrobacterium colonization of cotyledon surfaces as well as infection of internal cells. GFP was first detected 7 h following inoculation of the cotyledon. By 36 h, GFP expression was very intense, and was limited to the adaxial surface of the cotyledon. Expression of GFP also served as a useful indicator of successful elimination of the bacterium from plant tissue following antibiotic treatment.

Sonication-assisted Agrobacterium-mediated transformation of the ACC gene to interfere the production of ethylene in spring Dendrobium cv. ‘Sanya’

“…However, our preliminary experiments demonstrated that ultrasound treatment and a subse quent immersion in A. tumefaciens suspension for inoculation was more effective than treatment with ultrasound concurrently with A. tumefaciens inocula tion.. We suggest that ultrasound treatment forms microwounds on the surface of PLBs and facilitates Agrobacterium infection. The ultrasound treatment was successfully employed to enhance transformation efficiency in several other plant species [7][8][9][10][11][12]. The impact of different ultrasonic power on transgenic 'Sanya' requires further study.…”

“…Pathak and Hamzah [6] found that in chickpeas the transformation efficiency was more than twice higher with SAAT treatment than at simple Agrobacterium transformation without soni cation. Subsequently, the ultrasound treatment was successfully employed to enhance transformation effi ciency in several plant species, both by direct DNA transfer to protoplasts and tissue cultures and by co cultivation with vector bacteria [7][8][9][10][11][12].…”

Improving Infection Efficiency of Agrobacterium to Immature Cotyledon Explants of Japanese Apricot (Prunus mume) by Sonication Treatment

“…They found that sonication treatment could increase the transient expression of GUS by 100-to 1400-fold in several different plant species. Subsequently, they reported another study on the dynamics and optimization of Agrobacterium colonization and infection in the transformation of soybean cotyledons using sonication treatment and a GFP reporter (Finer and Finer, 2000). They described that GFP provided a quick, non-destructive method to evaluate, in real time, Agrobacterium colonization of cotyledon surfaces as well as infection of internal cells, and that sonication produced extensive microwounds on the surface of plant tissue, allowing increased access by the bacterium to plant cells.…”

Soybean: Plant Manipulation to Agrobacterium Mediated Transformation

“…They also reported that inoculum OD 600 1.0 gave better transient expression but no interaction was found between SAAT, Agrobacterium strain and soybean genotype. Micro-wound in plant tissues due to SAAT treatment release compounds that facilitate growth and accumulation of bacteria under aerobic conditions so facilitate transformation efficiency (Finer & Finer, 2000). However, longer sonication time may damage plant tissue (Santarem et al 1998).…”