In vitro direct plant regeneration and Agrobacterium‐mediated transformation of lucerne (Medicago sativa L.). (27th October 2012)

• Record Type: Journal Article
• Authors: Kumar, S.; Tiwari, R.; Chandra, A.; Sharma, A.; Bhatnagar, R. K.
• Is Part Of: Grass and forage science. Volume 68:Number 3(2013:Sep.)
• Page Start: 459
• Page End: 468
• Publication Date: 2012-10-27
• Subjects: Grasses -- Periodicals; Forage plants -- Periodicals; Grasslands -- Periodicals; 633.2
• DOI: 10.1111/gfs.12009
• Languages: English
• ISSNs: 0142-5242

Abstract

<abstract abstract-type="main" xml:lang="en" id="gfs12009-abs-0001"> <title>Abstract</title> <p> <italic>In vitro</italic> direct plant regeneration of lucerne was achieved by simultaneous application of thidiazuron (TDZ) and 6‐benzyladenine (BA) in Murashige and Skoog (MS) medium. Seedlings were germinated and grown for 6 d on growth regulator–containing MS medium. The shoot tip, consisting of the apical meristem along with parts of the cotyledonary leaves and hypocotyl, was then cultured on a medium containing the growth regulator(s). Adventitious budding of the shoot tip was promoted synergistically by treatment with TDZ and BA, and a maximum of thirty‐five shoots per explant was obtained on a medium supplemented with 2 mg L<sup>−1</sup> TDZ and 1 mg L<sup>−1</sup> BA. Plant regeneration frequency varied from 67 to 93%, and five Indian lucerne cultivars responded well to the regeneration protocol. The <italic>Agrobacterium</italic>‐mediated transformation frequency from co‐cultivated explants was 13% following multiple shoot induction. Southern analysis of the T<sub>0</sub> plants and T<sub>1</sub> progenies confirmed stable inheritance of the <italic>hpt</italic> marker gene. <italic>Agrobacterium</italic> infection of the explant caused a significant reduction in the plant regeneration frequency (23%) and the number of shoots induced (11) when compared with uninfected explants. A single shoot tip provided sufficient material to regenerate and establish twenty‐seven lucerne plants, whereas only nine plants could be regenerated from an <italic>Agrobacterium</italic> co‐cultivated explant. This transformation protocol could represent a valuable improvement over existing ones for lucerne.</p> </abstract>