Optimization of tissue culture and transformation of construct contain GDP-mannose-3´, 5´-epimerase (GME) gene from actinidia deliciosa L. into Lactuca sativa L

Lettuce is considered as a model plant for biotechnology because of its compatibility with stable genetic transformation and tissue culture. GDP-mannose-3’,5’-epimerase (GME) is one of the key genes in ascorbic acid biosynthesis pathway in plants. The present study aims to transfer GME gene from Actinidia deliciosa L. into Lactuca sativa L. 

To investigate callus induction rate using the effects of explant (cotyledon and true leaves) and 16 plant growth regulator combination including concentrations of 0.02, 0.04, 0.05, and 0.1 mg/l NAA and 0.1, 0.2, 0.4, and 0.6 mg/l BAP, and also direct regeneration rate using the effects of explant (cotyledon and true leaves) and 6 plant growth regulator combination including concentrations of 0.02 and 0.05 mg/l NAA and 0.2, 0.4, and 0.6 mg/l BAP, a factorial experiment based on completely randomized design with three replications was conducted. In order to transform GME into Lactuca sativa L. using L. sativa L. cv. Setareh and Agrobacterium tumefaciens strain (C58) on two types of explants (cotyledon and true leaves), a factorial experiment with three replications and 2 min and 8 min inoculation was done.

The results revealed that the highest percentage of callus induction and indirect regeneration (100%) were observed on leaf and cotyledon explants and MS medium containing 0.1 mg/l BAP and 0.04 mg/l NAA. The results also confirmed the presence of pBI121+GME in transgenic plants.

The explant true leaves and 2 min inoculation (with 18 percent transformation ratio) were more suitable for transformation.