Shoot regeneration from cultured leaf explants of Lycium barbarum and Agrobacterium-mediated transformation

A method for fast plant regeneration via organogenesis directly from Lycium barbarian leaf explants has been developed. The key factor for shoot regeneration was the presence of benzyladenine (BA) in the medium. NAA could only induce root formation and explant callusing. Murashige and Skoog (MS) medium supplemented with 2 mg/l BA and 0.5 mg/l NAA is the most efficient condition for shoot formation, with up to 92.6% shoot regeneration and no callus formation. All adventitious shoots cultured on MS medium supplemented with I mg/l iAA formed an extensive root system. Regenerated plants were morphologically normal and were also proved to be diploid (2n = 24). Using the optimized regeneration system, the genetic transformation of L barbarum was carried out mediated by Agrobacterium tumefaciens EHA101 (pIG 121 Hm). 11.8% leaf explants produced kanamycin-resistant shoots after infection by A. tumefaciens. The putative transgenic nature of plants was confirmed by GUS assay and PCR analysis. Expression of the nptII gene in the regenerated plants was also detected by observing the callus formation by leaf pieces on MS medium containing 0.2 mg/l 2,4-D and 0-100 mg/l kanamycin.