Abstract: 【Objective】The aims of this study are 1) to analyze the role of ath-miR169d in the progress of leaf development in Arabidopsis, 2) to illuminate the mechanisms of molecular regulation mediated by ath-miR169d, and 3) to provide a new insight into improving the photosynthetic performance and the genetic engineering of vegetables biomass and crop productivity.【Method】Ath-miR169d over-expression transgenic plants, STTM ath-miR169d knockdown transgenic plants, and WT plants were grown in a controlled culture room at 22℃ with a relative humidity of 60% and 16 h/8 h photoperiod. Number and size of rosette leaf were measured; pmiR169d::GUS vector were constructed and transformed into Agrobacterium tumefaciens EHA105, and then infected the buds of wildtype Arabidopsis (Columbia, Col-0), the pmiR169d::GUS transgenic Arabidopsis plants were used to observe the expression profile of ath-miR169d in different organs and tissues by GUS staining method. Using 8-week-old seedlings of ath-miR169d over-expressing transgenic Arabidopsis thaliana and wild type plants as materials, morphology of leaf epidemic cells were observed by scanning electron microscope (SEM). Whole shoots were harvested from 4-week-old seedlings of ath-miR169d over-expressing transgenic A. thaliana and wild type plants, subsequently the total endogenous IAA were detected and quantified as methyl esters by gas chromatography-mass spectroscopy (GC-MS). The mRNA expression profiles were detected by microarray analysis to screen differentially expressed genes. The expression of key genes in plant auxin signal pathway were verified by real-time fluorescent quantitative PCR (RT-qPCR). 【Result】In contrast to WT, ath-miR169d overexpression plants exhibited fewer and smaller rosettes, STTM ath-miR169d knockdown plants showed more and larger rosettes. Moreover, STTM ath-miR169d plants could generate new rosette leaves incessantly even after bolting and seeds harvest, whereas the leaves of ath-miR169d overexpression and WT plants generally decayed after harvest. The epidermal cells in the leaves of ath-miR169d over-expression plants were smaller than that in the WT leaves by SEM detection. Ath-miR169d was highly expressed in SAM and leaf vasculature, and expression profile was stronger in new leaves than in old ones. Endogenous IAA content reduced by 38.6% in ath-miR169d over-expressing plants compared to wild-type plants, showing Ath-miR169d was potentially involved in auxin signal pathway by microarray analysis. The auxin biosynthesis gene YUC2 and transporter gene PIN1 were downregulated in ath-miR169d over-expressing plants whereas, auxin response factors 1 and 2 (ARF1 and ARF2) genes were upregulated. Expression profile of YUC2, PIN1, ARF1 and ARF2 in STTM miR169d plantswere opposite to over-expressing plants. 【Conclusion】Results of the experiment showed that the leaf development was regulated by ath-miR169d through the auxin-signaling pathway. Number and size of rosette could be alerted by ath-miR169d expression level, and further affected biomass of the whole plant.

Key words: Arabidopsis, miR169, leaf, development, auxin signaling pathway