Biolistic transformation of broccoli (<i>Brassica oleracea</i>var.<i>italica</i>) for transient expression of the b-glucuronidase gene

Puddephat, I. J.; Thompson, Nancy; Robinson, Helen T.; Sandhu, P.; Henderson, Jeffrey

doi:10.1080/14620316.1999.11511178

Cited by 13 publications

(7 citation statements)

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“…Mukhopadhyay et al, 1991;Xue et al, 1997) or mesophyll cells (Radchuk et al, 2002;Nugent et al, 2006). Apart from transient expression data in broccoli (Puddephat et al, 1999) there are no reports of nuclear transformants of Brassica oleracea via biolistics. Interestingly, biolistics has been used to produce plastid transformants of Brassica napus (Hou et al, 2003) and B. oleracea (Liu et al, 2007) from seedling and leaf explants, respectively.…”

Section: Introductionmentioning

confidence: 99%

Regeneration from leaf explants and protoplasts of Brassica oleracea var. botrytis (cauliflower)

Chikkala

Nugent

Dix

et al. 2009

Scientia Horticulturae

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Section: Introductionmentioning

confidence: 99%

Regeneration from leaf explants and protoplasts of Brassica oleracea var. botrytis (cauliflower)

Chikkala

Nugent

Dix

et al. 2009

Scientia Horticulturae

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“…The particle bombardments using variable parameters have been detailed by Heiser, (1995). In broccoli, consistently high rate of transformation was obtained using M17 Tungsten particles fired at 900 psi with a gap distance of 20 mm and target distance of 55 mm (Puddephat et al, 1999).…”

Section: Biolistic Transformationmentioning

confidence: 94%

“…This is applicable to a wide range of plant genotypes, especially those in which embryogenic cell cultures are available (Kikkert et al, 2005). This has already been used in other vegetables like broccoli (Puddephat et al, 1999), garlic (Sawahel, 2002) and potato (Craig et al, 2005).…”

Section: Molecular Analysismentioning

confidence: 99%

“…Agrobactrium mediated transformation of Cry1Ac gene in eggplant, has been reported (Franklin and Sita, 2003;Soniya et al, 2005), but Biolistic mediated is not there in literature. This method has been proved useful to deliver DNA into cells of wide range of plants (Klein et al, 1988) and used successfully vegetables like broccoli (Puddephat et al, 1999), garlic (Sawahel, 2002) and potato (Craig et al, 2005). Therefore, experiment was conducted on standardization of regeneration protocol, transformation of Cry1Ac gene using Biolistic gun and molecular evaluation of the putative transgenic plant.…”

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confidence: 99%

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Biolistic transformation ofcry 1Ac gene in eggplant (Solanum melongenaL.)

Sidhu

Dhatt²,

Sandhu³

et al. 2014

Intern. Jour. of Agricul., Environ. and Biotech.

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Among different explants, cotyledon showed highest regeneration and number of buds per explant on all media combinations. However, highest regeneration potential was observed on MS fortified with 3.0 mgl -1 BAP (80.36%), but number of buds per explant were highest (21.76) on MS media supplemented with 2.5 mgl -1 BAP+ 1.0 mgl -1 Kin. Plant regeneration of cotyledons was completely checked with a selection pressure of 15ppm to 25ppm hygromycin for untranformed explants. But the selection efficacy of transformed tissue was 19.56% with 20ppm and 66.66% with 15 ppm hygromycin. GUS gene was expressed in 76.92% bombarded explants with 2 to 7 GUS spots per explant confirming the presence of transformed tissue. GUS gene was also visualized in 66.66% regenerated shoots from bombarded tissue. PCR analysis of putative transgenic eggplant plants with the specific primers confirmed the presence of Cry1Ac gene with 1.2 KB product size (falling between 1KB and 2KB bands of 10KB DNA ladder) in two of the four plants. Highlights• Cotyledon of genotype BL-5 had highest regeneration potential and number of buds per explant on MS media supplemented with 2.5 mgl -1 BAP+ 1.0 mgl -1 Kin.• Plant regeneration of cotyledons was completely checked with a selection pressure of 15ppm to 25ppm hygromycin for untranformed explants.• The selection efficacy of transformed tissue was 19.56% with 20ppm and 66.66% with 15 ppm hygromycin.• GUS gene was expressed in bombarded explants as well as in regenerated putative transgenic explants.• PCR analysis of putative transgenic eggplant plants with the specific primers confirmed the presence of Cry1Ac gene.

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“…However, apart from transient reporter gene expression in broccoli [34], biolistics has not produced transgenic vegetable Brassicas. Furthermore, although biolistics is the most efficient method for plastid transformation, a protoplast system while being more labour-intensive, is cheaper.…”

Section: Introductionmentioning

confidence: 99%

Nuclear and plastid transformation of Brassica oleracea var. botrytis (cauliflower) using PEG-mediated uptake of DNA into protoplasts

et al. 2006

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Most plastid transformation studies with tobacco, and all reports for other species (except tomato [G.D. Nugent, M. ten Have, A. van der Gulik, P.J. Dix, B.A. Uijtewaal, A.P. Mordhorst, Plastid transformants of tomato selected using mutations affecting ribosome structure. Plant Cell Rep. 24 (2005) 341-349]), have used biolistics for plastid transformation. However, nuclear transformation via biolistics has not been reported for any vegetable Brassica species so we used protoplast culture and PEG-mediated DNA uptake, to examine both nuclear and plastid transformation of cauliflower, an important vegetable Brassica. A vector containing genes for hygromycin resistance and b-glucuronidase activity (pGUS-HYG) was used for nuclear transformation, while plastid transformation utilised a vector (pZB1) containing accD-rbcL plastome targeting regions cloned from Brassica napus (oil seed rape), and the selectable marker gene aadA, conferring resistance to spectinomycin. Protoplasts were embedded in agarose and selected on media containing hygromycin or spectinomycin. From five experiments, a single plastid transformant of the commercial cultivar Thalassa was obtained, whereas nuclear transformants were obtained at an absolute transformation frequency up to 1.3 Â 10 À5. No spontaneous spectinomycin resistant mutants were observed in any plastid transformation experiments. PCR and Southern blot analysis confirmed the transgenic status of plants regenerated from the protoplast-derived calli. #

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Biolistic transformation of broccoli (Brassica oleraceavar.italica) for transient expression of the b-glucuronidase gene

Cited by 13 publications

References 0 publications

Regeneration from leaf explants and protoplasts of Brassica oleracea var. botrytis (cauliflower)

Regeneration from leaf explants and protoplasts of Brassica oleracea var. botrytis (cauliflower)

Biolistic transformation ofcry 1Ac gene in eggplant (Solanum melongenaL.)

Nuclear and plastid transformation of Brassica oleracea var. botrytis (cauliflower) using PEG-mediated uptake of DNA into protoplasts

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