Agrobacterium tumefaciens-mediated genetic transformation of barley (Hordeum vulgare L.)

Shrawat, Ashok K.; Becker, Dirk; Lörz, Horst

doi:10.1016/j.plantsci.2006.09.005

Cited by 68 publications

(43 citation statements)

References 38 publications

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“…Agrobacterium-mediated transformation has several advantages such as high transformation efficiency compared to protoplast transfer (Rao et al 2009), the ability to transfer large pieces of DNA, minimal re-arrangement of transferred DNA, and characteristic insertion into the recipient genome of a discrete segment of DNA at a low copy number (Kumar et al 2005;Tyagi et al 2007) compared to particle bombardment approach (Shrawat et al 2006;Rao et al 2009). Efficiency of Agrobacterium-mediated transformation and delivery of T-DNA into plant cells is influenced by several parameters including explant selection (Owens & Cress 1985;Hiei et al 1994;Ishida et al 1996).…”

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confidence: 99%

The effect of different antibiotics on the elimination of Agrobacterium and high frequency Agrobacterium-mediated transformation of indica rice (Oryza sativa L.)

Priya¹,

Subramanian²,

Manikandan³

2012

CZECH J GENET PLANT

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Abstract:We report here the suitable explant with high efficiency of transformation and the positive effects of timentin over other antibiotics like carbenicillin and cefotaxime on the elimination of Agrobacterium tumefaciens during the genetic transformation of popular indica rice (Oryza sativa L.). The tissues assayed were embryogenic calli, embryos with endosperm contamination, intact seeds, leaf blades, leaf bases and coleoptiles. The frequency of transient β-glucuronidase (GUS) expression as revealed by histochemical assay was 90% for embryogenic calli, which was the highest among the explants used. On the basis of disc-diffusion assay, the maximum zone of inhibition (29 mm) at 250 mg/l was obtained for timentin. In tissue culture conditions the frequency of Agrobacterium recurrence after 20 days of infection was minimum (2.3%) at 200 mg/l of timentin. At 250 mg/l of timentin there was no Agrobacterium growth, besides, there were no negative effects on the callus growth unlike other antibiotics, hence it was selected as the optimum concentration for high frequency callus proliferation and regeneration. The effect of the parameters evaluated was determined by the callus proliferation during selection, reduction in browning, transient GUS expression and stable transformation efficiency (23.3%). The resultant plants were stable transformants as confirmed by a molecular analysis of the gus and hpt genes. The developed transformation protocol will be very helpful for the information on indica rice cultivars in general and on IR 64 in particular.

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confidence: 99%

The effect of different antibiotics on the elimination of Agrobacterium and high frequency Agrobacterium-mediated transformation of indica rice (Oryza sativa L.)

Priya¹,

Subramanian²,

Manikandan³

2012

CZECH J GENET PLANT

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“…Although relatively few transgenes were confirmed in explants (17 of 161, or 10.5%), this ratio of transformation is higher than that which is commonly observed using the Agrobacterium system and in transformation rates reported for Begonia x hiemalis Fotsch (0-3%) (Kishimoto et al, 2002); Hordeum vulgare (0-7%) (Shrawat et al, 2007), and an Vigna unguiculata (1%) (Chaudhury et al, 2007). However, considering the influence of ultrasound as well as time of contact with the bacteria, there was a significant increase in the number of transformed plants.…”

Section: Discussionmentioning

confidence: 53%

Improving plant transformation using Agrobacterium tumefaciens

Neto¹,

Oliveira²,

Lourenço³

et al. 2015

Genet. Mol. Res.

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ABSTRACT. Here, we report a quick and low-cost method to improve plant transformation using Agrobacterium tumefaciens. This method involves the use of physical wounding, ultrasound, and an increase in exposure time to the bacteria. We show how the transformation rate increased from 0 to 14% when an ultrasound pulse of 10 s was used in conjunction with 96 h of bacterial exposure in Eclipta alba explants.

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“…The synergistic effects of 2,4-D and dicamba might have also contributed to the production of high quality embryogenic spherical nodular calli from cocultured calli. The effect of a combination of two auxins on production of high quality embryogenic callus induction has also been reported in wheat (Afshar-Sterle 1996) and barley (Shrawat et al 2007). In addition, early differentiation of these embryogenic nodular calli was overcome by adding ABA and a combination of Dicamba and 2,4-D. Embryogenic units between 3-5 mm in size, which were compact and spherically shaped with a yellowish rough surface, produced the maximum numbers of resistant cell lines after co-cultivation with Agrobacterium and consequently the highest frequency of transgenic events.…”

Section: Discussionmentioning

confidence: 83%

“…Addition of these amino acids in the medium gave a rapid increase in the production of highly embryogenic spherical nodules. The combination of these amino acids has been reported in promoting the production of somatic embryos and green regenerants in wheat (Afshar-Sterle 1996) and barley (Shrawat et al 2007). The synergistic effects of 2,4-D and dicamba might have also contributed to the production of high quality embryogenic spherical nodular calli from cocultured calli.…”

Section: Discussionmentioning

confidence: 99%

A high-throughput Agrobacterium tumefaciens-mediated transformation system for molecular breeding and functional genomics of rice (Oryza sativa L.)

Shrawat

Good

2010

Plant Biotechnology

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Agrobacterium tumefaciens-mediated transformation is the preferred method for genetic transformation of rice. Here we provide a comprehensive and high-through-put protocol for Agrobacterium-mediated transformation of rice for generating the large numbers of transgenic plants that are required for functional analysis and for evaluating traits of agronomic importance. In a project designed to produce nitrogen use efficient rice plants, we refined/improved several factors including optimization of conditions for inducing vir genes prior to transformation, infection and co-cultivation medium for better interaction of target tissues with Agrobacterium. These optimizations supported not only the survival of co-cultured calli in high frequency, but also the production of multiple resistant cell lines per co-cultured embryogenic and nodular callus. In addition, improvements in plant tissue culture, selection and regeneration media has enabled us to produce large numbers of transgenic rice plants containing genes of agronomical importance. Partial desiccation and ABA treatment to hygromycin-resistant callus tissue significantly (PϽ0.05) enhanced both regeneration frequency and regeneration of transgenic plantlets per callus tissue. Regeneration frequency was further improved by optimizing the concentration of copper sulphate in the regeneration medium. The majority of the transgenic plants obtained using this improved protocol displayed a normal phenotype and the gene of interest was inherited by the progeny in a Mendelian fashion. Homozygous plants of selected lines showed stable phenotypes both in soil and hydroponic conditions even after five generations. Availability of such a high-throughput Agrobacterium-mediated transformation system will improve future opportunities for rice genetics and functional genomics study.

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Agrobacterium tumefaciens-mediated genetic transformation of barley (Hordeum vulgare L.)

Cited by 68 publications

References 38 publications

The effect of different antibiotics on the elimination of Agrobacterium and high frequency Agrobacterium-mediated transformation of indica rice (Oryza sativa L.)

The effect of different antibiotics on the elimination of Agrobacterium and high frequency Agrobacterium-mediated transformation of indica rice (Oryza sativa L.)

Improving plant transformation using Agrobacterium tumefaciens

A high-throughput Agrobacterium tumefaciens-mediated transformation system for molecular breeding and functional genomics of rice (Oryza sativa L.)

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