Development of a rapid, highly efficient system of organogenesis in cowpea Vigna unguiculata (L.) Walp

Raveendar, Sebastin; Premkumar, A.; Sasikumar, S.; Ignacimuthu, S.; Agastian, Paul

doi:10.1016/j.sajb.2008.05.009

Cited by 18 publications

(14 citation statements)

References 22 publications

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“…However, several authors have reported direct plant regeneration from both hypocotyls (Annapurna and Rathore 2010;Nagori and Purohit 2004;Shang et al 2006;Singh et al 2002) and cotyledons (Du and Pijut 2008;Raveendar et al 2009). The data presented in this study clearly demonstrate that high-frequency adventitious shoot regeneration is possible from both hypocotyl and cotyledon segments of L. frutescens, also a woody species.…”

Section: Discussionmentioning

confidence: 99%

“…Following 4-weeks of culture on plant growth regulator-supplemented media, the adventitious shoots in this study continued to proliferate, showing no signs of senescence or necrosis. Furthermore, the shoots did not require additional transfer to shoot multiplication or elongation medium, which is a standard procedure (Chitra and Padmaja 2005;Kumar et al 2005;Quintero-Jiménez et al 2010;Raveendar et al 2009;Zhang et al 2008), and were subsequently transferred directly onto rooting medium, thereby eliminating the need for an additional multiplication or elongation phase. In terms of shoot production time, this technique is a more efficient and rapid method of deriving shoots than the indirect organogenesis protocol or the axillary bud culture previously reported (Dewir et al 2010;Shaik et al 2010b).…”

Section: Discussionmentioning

confidence: 99%

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Cytokinin-induced organogenesis in Lessertia (Sutherlandia) frutescens L. using hypocotyl and cotyledon explants affects yields of l-canavanine in shoots

Shaik

Singh

Nicholas

2010

Plant Cell Tiss Organ Cult

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A simple protocol for direct shoot organogenesis and plant regeneration in Lessertia frutescens using hypocotyl and cotyledon segments is reported. L-canavanine content in the derived shoots is also quantified. Media containing different concentrations and combinations of the cytokinins kinetin (K) and benzyladenine (BA) were tested for shoot induction potential. The best shoot regeneration rate (83%) was obtained from hypocotyl segments cultured in Murashige and Skoog (MS) medium supplemented with 1 mg l -1 K; these hypocotyls also produced the largest number of shoots per explant (3.5) and the longest shoots per explant (13.3 mm). The best shoot regeneration rate (46%) using cotyledons as explant material was obtained in MS medium supplemented with 1 mg l -1 K and 1 mg l -1 BA or with 5 mg l -1 K and 0.5 mg l -1 BA. The highest number of cotyledon-derived shoots (1.5) was obtained in MS medium containing 2 mg l -1 K and 0.5 mg l -1 BA, and the longest cotyledon-derived shoots (6.1 mm) were obtained in MS medium containing 1 mg l -1 K and 0.5 mg l -1 BA. Shoots derived from hypocotyls cultured on media containing 1 mg l -1 K contained the highest quantity of L-canavanine (1.42 mg g -1 ) relative to the control (0.52 mg g -1 ). Shoots derived from cotyledons cultured on media containing 2 mg l -1 K contained the highest quantity of L-canavanine (2.07 mg g -1 ) compared to the control. Scanning electron microscopy revealed that shoots regenerated directly from the wounded epidermal tissue, although callus formation was observed in most cultures. Young shoot clusters proliferated into healthy adventitious shoots that were subsequently transferred directly onto rooting medium (MS medium containing 4 mg l -1 indole-3-butyric acid), eliminating the need for an additional multiplication or elongation phase. The in vitro plants were successfully acclimatized in a growth chamber, achieving an 85% survival rate.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Cytokinin-induced organogenesis in Lessertia (Sutherlandia) frutescens L. using hypocotyl and cotyledon explants affects yields of l-canavanine in shoots

Shaik

Singh

Nicholas

2010

Plant Cell Tiss Organ Cult

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“…De novo shoot organogenesis using EA as explants A rapid, efficient and reproducible regeneration system is a prerequisite for establishment of an efficient cowpea genetic transformation system. Although several studies of in vitro regeneration of cowpea based on organogenesis have been reported (Aasim et al, 2010;Abdu Sani et al, 2015;Mamadou et al, 2008;Manman et al, 2013;Odutayo et al, 2005;Raveendar et al, 2009;Sani et al, 2018;Tie et al, 2013;Yusuf et al, 2008), an efficient cowpea regeneration system that enables highly efficient transformation is still lacking (Manman et al, 2013). Soybean transformation based on the preexisting meristems of EA explants has been well established and provides a reliable and highly efficient mean for introducing transgenes (Aragão et al, 2000;Liu et al, 2004;Turlapati et al, 2008).…”

Section: Resultsmentioning

confidence: 99%

Developing a rapid and highly efficient cowpea regeneration and transformation system using embryonic axis explants

Che

Chang

Simon

et al. 2019

Preprint

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Cowpea is one of the most important legume crops planted worldwide, especially in Sub-Saharan Africa and Asia. Despite decades of effort, genetic engineering of cowpea is still challenging due to inefficient in vitro shoot regeneration, Agrobacterium-mediated T-DNA delivery and transgenic selection. Here, we report a rapid and highly efficient cowpea transformation system using embryonic axis explants isolated from imbibed mature seeds. We found that removal of the shoot apical meristem by cutting through the middle of the epicotyl stimulated direct multiple shoot organogenesis from the cotyledonary node tissue.Furthermore, the application of a ternary transformation vector system using an optimized pVIR accessory plasmid provided high levels of Agrobacterium-mediated gene delivery. The utilization of spectinomycin as the selection agent enabled more efficient transgenic selection and plant recovery. Transgenic cowpea shoots developed exclusively from the cotyledonary nodes at high frequencies of 4.5 to 37% across a wide range of cowpea genotypes. We believe that the transformation principles established in this study could also be applied to other legumes to increase transformation efficiencies.

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“…Establishment of an efficient and reproducible transformation system is based on efficient plant tissue culture using various explants (Muthukumar et al, 1995;Prem Anand et al, 2001;Choi et al, 2003;Ramakrishnan et al, 2005;Chaudhury et al, 2007;Aasim et al, 2012) and growth regulators used for shoot regeneration (Muthukumar et al, 1995;Monti et al, 1997;Pellegrineschi, 1997;Brar et al, 1999;Machuka et al, 2000;Obembe et al, 2000;Raveendar et al, 2009). No callusing was recorded on regenerating explants, in contradiction to the findings of Choi et al (2003).…”

Section: Discussionmentioning

confidence: 99%

Production of herbicide-resistant cowpea (Vigna unguiculata L.) transformed with the bar gene

Aasım¹,

Khawar²,

Özcan³

2013

Turk J Biol

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Plant genetic engineering has opened new avenues to modify crops and has provided a powerful tool for crop improvement.\ud The present study reports the development of regeneration and genetic transformation protocol for the Turkish cowpea cultivar Akkiz.\ud The immature cotyledons were cultured on Murashige and Skoog (MS) medium containing 0.25–0.75 mg L–1 6-benzylaminopurine with\ud or without 0.25 mg L–1 α-naphthalene acetic acid. Shoot regeneration varied 44.4%–83.3% with 2.1–5.0 shoots per explant. Regenerated\ud shoots were rooted in MS medium containing 0.50 mg L–1 indole-3-butyric acid and acclimatized in the greenhouse, where they flowered\ud and set seeds. Immature cotyledons were transformed with Agrobacterium tumefaciens strain LBA4404 harboring the recombinant\ud binary vector pRGG containing an herbicide tolerance gene (bar) along with a uida (GUS) gene under 35S promoter. Phosphinothricin\ud was used as a selectable marker at a concentration of 2.5 mg L–1. Putative transformants were screened by the histochemical GUS assay.\ud Furthermore, molecular analysis revealed the presence of the introduced gene in the genome of cowpea cultivar Akkiz. The selected\ud transgenic plants showed a resistance to Basta® nonselective herbicide at up to 10 mL L–1 of water. Putative transgenic plants retained\ud their pigmentation and continued to grow in the greenhouse

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Development of a rapid, highly efficient system of organogenesis in cowpea Vigna unguiculata (L.) Walp

Cited by 18 publications

References 22 publications

Cytokinin-induced organogenesis in Lessertia (Sutherlandia) frutescens L. using hypocotyl and cotyledon explants affects yields of l-canavanine in shoots

Cytokinin-induced organogenesis in Lessertia (Sutherlandia) frutescens L. using hypocotyl and cotyledon explants affects yields of l-canavanine in shoots

Developing a rapid and highly efficient cowpea regeneration and transformation system using embryonic axis explants

Production of herbicide-resistant cowpea (Vigna unguiculata L.) transformed with the bar gene

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