Analysis Of Molecular Basis In Shoot Regeneration Between Lateral Root Primordium And Callus Pathways

Plants can regenerate shoot apical meristems by producing calli under appropriate in vitro culture conditions,and subsequently regenerate shoot and the whole plant.Recent studies have found that treatment with appropriate concentration of hormone can directly transdifferentiate the lateral root primordia into shoot apical meristem without callus formation.However,the similarity and differences between the mechanisms underlying the two pathways remains unknown.In this study,a shoot regeneration system was established wherein the explants were cultured in the medium containing auxin NAA and subsequently cytokinin 2-iP,and regenerated shoots from callus and lateral root primordium at the same time.The difference between the mechanisms of the two shoot regeneration pathways were examined.The main results are as follows:Morphological observation showed that in the process of shoot regeneration from lateral root primordium,the lateral root primordium developed in the early stage,the converting organs formed in the middle stage,and shoot meristem developed in the late stage.In the process of shoot regeneration from callus,the callus developed in the early stage,the converting organ formed in the middle stage and the shoot meristem formed in the late stage.The transcriptometic results showed that the genes related to root development were expressed in both pathways,indicating that the callus had the similar properties with root primordium,which was consistent with previous findings.Further analysis showed that the genes involved in plant hormone signal transduction and synthesis had significant differences between two pathways.The transcription levels of auxin synthetic gene YUCCA1(YUC1),AUXIN RESPONSE FACTOR 5(ARF5)which encodes auxin signal component and WUSCHEL-RELATED HOMEOBOX 5(WOX5),a key gene for stem cell maintenance,were significantly higher in callus-derived shoots than those of lateral root primordial-derived ones.But the expression of YUC2 showed the opposite expression pattern.Moreover,although YUC1 and ARF5 showed similar expression patterns between two pathways,their expression signals were significantly stronger in callus-derived shoots at the quiescent period.Auxin response signal DR5 also showed similar distribution patterns.The key regulatory gene for stem cell maintenance WUSCHEL(WUS),cytokinin synthesis related gene ISOPENTENYLTRANSFERASE 5(IPT5)and Arabidopsis Response Regulator 10(ARR10)encoding cytokinin signal component exhibited similar expression patterns in both pathways.In the arf5 and yuc1 mutants,frequencies of shoot regenerated from callus were significantly decreased,while that derived from lateral root primordium transdifferentiation did not changed obviously,indicating that YUC1 and ARF5 specifically regulated shoot regenerated from callus.The results of this study provide new information for understanding the regulatory mechanism of shoot meristem regeneration and cell fate transformation,and lay a foundation for establishing new systems for plant in vitro propagation and genetic transformation.