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

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Summary

Introduction

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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