In vitro flowering and viable seed setting of transgenic lettuce cultures

Abstract

Production of transgenic lettuce seeds via in vitro flowering and fruit setting is reported here. Six days old cotyledons were co-cultivated with Agrobacterium tumefaciens strain EHA105 harbouring the binary vector pCAMBIA2301 carrying the reporter gene α-glucuronidase intron (GUS-INT) and the marker gene neomycin phosphotransferase (NPTII). Transgenic calluses and shoot buds were induced on MS medium augmented with 0.1 mg l⁻¹ benzyladenine (BA), 0.1 mg l⁻¹ α-naphtaleneacetic acid (NAA), 100 mg l⁻¹ kanamycin and 500 mg l⁻¹ timentin (ticarcillin clavulanate). After transferring the cultures onto MS basal medium augmented or not with kanamycin, in vitro flower induction was observed within 90 days and eventually matured and produced seed pods. Pollen grains obtained from flowers induced on basal medium without kanamycin were heterogeneous containing both GUS positive and negative pollen grains. In the corresponding seed population, a Mendelian ratio (3 : 1) of gusA transgene segregation was observed. On the other hand, flowers that were induced on basal medium under kanamycin selection, all the pollen grains were GUS positives.