A tobacco microsomal ω-3 fatty acid desaturase gene increases the linolenic acid content in transgenic sweet potato (Ipomoea batatas)

As a first step to the establishment of a genetic transformation protocol for olive somatic embryos obtained from the seeds of cv. 'Picual', the efficiencies of different aminoglycoside antibiotics as selective agents to be used with the nptII marker gene, and the particle bombardment technique for transient transformation have been evaluated. Among the three antibiotics tested, paromomycin and kanamycin showed a similar inhibitory effect and, at 200 mg l -1 , both of them impaired callus growth after 8 weeks of culture. However, when isolated embryos were cultured in the presence of these antibiotics, a 20% of the embryos still remained viable at 400 mg l -1 . Neomycin was discarded as a selective agent since it showed only a moderate toxic effect. Contrary to solid medium, when olive callus was cultured in liquid medium supplemented with different paromomycin concentrations for 3 weeks, the callus growth was impaired at the lowest antibiotic concentration, 3 mg l -1 . Best conditions for transient transformation of olive callus using PDS-1000/He system were a 6 cm target distance and a 900 psi bombardment pressure. pCGUD1 plasmid, containing the gus gene under the control of sunflower ubiquitin promoter yielded a significantly higher number of gus expression areas per bombarded explant than pGUSINT or pJGUS5 plasmids, where the gus gene is driven by CaMV35S promoter or CaMV35S with enhancer, respectively. Almost 45% of bombarded explants showed gus expression 12 weeks after bombardment.

Section: Discussionmentioning

confidence: 83%

Studies on genetic transformation of olive (Olea europaea L.) somatic embryos: I. Evaluation of different aminoglycoside antibiotics for nptII selection; II. Transient transformation via particle bombardment

Pérez-Barranco

Torreblanca

Padilla

et al. 2009

“…Different explants such as leaves, petioles, stems, storage roots, and embryogenic calli have been used for A. tumefaciens-mediated transformation of sweetpotato and stable transgenic sweetpotato plants have also been reported, but in most cases only a low transformation efficiency was obtained (Newell et al 1995;Gama et al 1996;Morán et al 1998;Otani et al 1998Otani et al , 2001Otani et al , 2003Cipriani et al 1999Cipriani et al , 2001Wakita et al 2001;Kimura et al 2001;Song et al 2004;Shimada et al 2006;Luo et al 2006). In comparison, apical meristem-derived embryogenic calli can give an improved transformation efficiency in sweetpotato (Gama et al 1996;Otani et al 1998), but they are not readily available target tissues for most of sweetpotato cultivars due to low frequencies of embryogenic callus formation in apical meristem cultures (Al-Mazrooei et al 1997;Liu et al 1997;Wang et al 1998).…”

Section: Introductionmentioning

confidence: 93%

Efficient Agrobacterium tumefaciens-mediated transformation using embryogenic suspension cultures in sweetpotato, Ipomoea batatas (L.) Lam.

Zhai

Wang

et al. 2007

Efficient Agrobacterium tumefaciens-mediated transformation using embryogenic suspension cultures in sweetpotato, Ipomoea batatas (L.) Lam.Abstract Efficient Agrobacterium tumefaciensmediated transformation was achieved using embryogenic suspension cultures of sweetpotato (Ipomoea batatas (L.) Lam.) cv. Lizixiang. Cell aggregates from embryogenic suspension cultures were cocultivated with the A. tumefaciens strain EHA105 harboring a binary vector pCAMBIA1301 with gusA and hygromycin phosphotransferase II gene (hpt II) genes. Selection culture was conducted using 25 mg l À1 hygromycin. A total of 2,218 plants were regenerated from the inoculated 1,776 cell aggregates via somatic embryogenesis. b-glucuronidase (GUS) assay and PCR, dot blot and Southern blot analyses of the regenerated plants randomly sampled showed that 90.37% of the regenerated plants were transgenic plants. The number of integrated T-DNA copies varied from 1 to 4. Transgenic plants, when transferred to soil in a greenhouse and a field, showed 100% survival. No morphological variations were observed in the ex vitro transgenic plants. These results exceed all transformation experiments reported so far in the literature in quantity of independent events per transformation experiment in sweetpotato.

“…Sweet potato transformation using A. tumefaciens has been attempted from various explant tissues (Newell et al 1995;Gama et al 1996;Morán et al 1998;Mitchell et al 1998;Otani et al 1998Otani et al , 2001Otani et al , 2003Cipriani et al 1999;Wakita et al 2001;Kimura et al 2001;Song et al 2004;Shimada et al 2006;Luo et al 2006), but most of these studies showed a genotype-dependent response along with low transformation and regeneration efficiency. An efficient and reliable transformation and regeneration system is a prerequisite to the successful application of the genetic engineering approach to a crop species.…”

Section: Introductionmentioning

confidence: 97%

“…Transgenic sweet potato lines with improved nutritional quality (López et al 1996;Wakita et al 2001) and resistance against various biotic (Newell et al 1995;Cipriani et al 2001;Okada et al 2001) and abiotic stresses (Lim et al 2007) have already been developed. Sweet potato transformation has been attempted by electroporation Mitchell et al 1998;Lawton et al 2000;Okada et al 2001), particle bombardment (Prakash and Varadarajan 1992), and Agrobacterium rhizogenes (Otani et al 1993).…”

Section: Introductionmentioning

confidence: 98%

Transgenic sweet potato expressing mammalian cytochrome P450

Anwar

Watanabe

2010

Sweet potato [Ipomoea batatas (L.) Lam] is considered to be recalcitrant to transformation and regeneration because of its genotype-dependent in vitro responses. The lack of a genotype-independent transformation and regeneration system limits biotechnological applications in this plant species. To establish a transformation system for a diverse group of sweet potato genotypes, we examined sweet potato regeneration after transformation in five cultivars. An Agrobacterium tumefaciens transformation system was used for the introduction of mammalian cytochrome P450 genes, which are capable of conferring herbicide tolerance. Among the different factors studied, including explant type, plasmid vectors, and auxin type in the initiation media, auxin type had the greatest effect on the regeneration response. Of the auxins tested, only 4-fluorophenoxyacetic acid (4FA) induced regeneration from all cultivars. In terms of the quality of calli, 4FA promoted the induction of type I calli, which were capable of somatic embryo formation, whereas type II calli fail to produce somatic embryos. The frequency of somatic embryo formation was also affected by the composition of the embryo-induction media. Transgenic plants were regenerated from all cultivars. The stable integration and expression of transgenes was confirmed by several approaches. This Agrobacterium-mediated transformation system should be applicable to a wide range of sweet potato cultivars.