Abstract
BackgroundLEAFY COTYLEDON 2 (LEC2) acts throughout embryo morphogenesis and maturation phase to maintain embryogenic identity. Our previous study stated that Arabidopsis thaliana LEC2 (AtLEC2) driven by glucocorticoid receptor-dexamethasone (GR-DEX) inducible system (AtLEC2-GR) triggers embryogenic callus formation in tobacco (Nicotiana tabacum).ResultsIn this study, the adenosine phosphate isopentenyltransferase genes AtIPT3, AtIPT7 and the tRNA isopentenyltransferase gene AtIPT9 were overexpressed in the AtLEC2-GR transgenic background. In the AtIPT7-OE AtLEC2-GR and AtIPT9-OE AtLEC2-GR seedlings, high-quality embryogenic callus was obtained under the DEX condition, and the shoot regeneration efficiency was 2 to 3.5 folds higher than AtLEC2-GR alone on hormone free medium without DEX. Transcriptome analyses showed that up-regulated BBM, L1L, ABI3, and FUS3 might function during embryogenic callus formation. However, at the shoot regeneration stage, BBM, L1L, ABI3, and FUS3 were down-regulated and Type-B ARRs were up-regulated, which might contribute to the increased shoot regeneration rate.ConclusionsA novel system for inducing shoot regeneration in tobacco has been developed using the GR-DEX system. Induced expression of AtLEC2 triggers embryogenic callus formation and overexpression of AtIPT7 or AtIPT9 improves shoot regeneration without exogenous cytokinin.
Highlights
LEAFY COTYLEDON 2 (LEC2) acts throughout embryo morphogenesis and maturation phase to maintain embryogenic identity
These results demonstrated that exogenous cytokinin could significantly promote the shoot regeneration efficiency of embryogenic callus from the AtLEC2GR transgenic line
Our results suggested that the up-regulation of gibberellic acids (GAs) biosynthesis enzyme genes might be another reason that caused the inhibition of embryogenic callus formation in the AtIPT3-OE Arabidopsis thaliana LEC2 (AtLEC2)-Glucocorticoid receptor (GR) seedlings
Summary
LEAFY COTYLEDON 2 (LEC2) acts throughout embryo morphogenesis and maturation phase to maintain embryogenic identity. Overexpression of a number of transcriptional factor genes can improve somatic embryogenesis and enhance plant regeneration, such as LEAFY COTYLEDON 1 (LEC1) [4], LEAFY COTYLEDON 2 (LEC2) [5, 6], WUSCHEL (WUS) [7], BABY BOOM (BBM) [8] and AGAMOUS-LIKE 15 (AGL15) [9]. LEC1/LEC1-LIKE (L1L) with three B3 domain protein genes ABSCISIC ACID (ABA)-INSENSITIVE3 (ABI3), FUSCUA3(FUS3) and LEC2 is referred as LAFL network (LEC1/L1L, ABI3, FUS3 and LEC2) [10]. This network functions redundantly throughout the early embryo developmental process, embryo maturation and dormancy in a dosedependent manner [11].
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