Cloning And Functional Identification Of SPL3,9,10 Genes In Betula Platyphylla

White birch(Betula platyphylla Suk.)is one of the important tree species in northern China,unisexual flowers,androgynous,has important research value.SPL(SQUAMOSA PROMOTER BINDING PROTEIN-LIKE)transcription factor is a plant-specific gene family,widely found in green plants,it plays an important role in regulating leaf development,flower and fruit development,fertility and so on.Firstly,through the information of genomic database,the ORF sequences of BpSPL3,BpSPL9 and BpSPL10 genes were cloned from birch,and the corresponding expression vectors and inhibitory expression vectors were constructed,which were named as:35S::BpSPL3-GFP,35S::BpSPL9-GFP,35S::BpSPL10-GFP,35S::BpSPL3-RNAi,35S::BpSPL9-RNAi and 35S::BpSPL10-RNAi.At the same time,the promoter sequence of the upstream 2000bp of the starting codon is cloned by the method of double enzyme cutting and the expression vector with GUS label is constructed,which is named BpSPL3::GOS,BpSPL9::GUS,BpSPL10::GUS in turn.The promoter of BpSPL3,BpSPL9 and BpSPL10 genes was predicted,and it was found that the promoter sequences of the above genes,in addition to the core promoter components,also contained light response elements,corresponding components of hormones,non-biological stress response elements and so on.By means of Agrobacterium-mediated method,the BpSPL3::GUS expression vector was transformed into birch and dipped in the stable genetic transformation to Arabidopsis,and the results showed that the BpSPL3 promoter had a high expression on the hairy body,stem,bud and root of the leaves in the birch,and the young leaf hairy body,the leaf edge of the old leaf in Arabidopsis,There was high GUS expression activity in lateral root and pistil stigma.The BpSPL3 gene specificity was found to be located in the nucleus by the method of sub-cell localization.Agrobacterium-mediated method for genetic transformation to obtain BpSPL3 gene transgenic strains.Through the analysis of transgenic strains,it was found that the root length of transgenic plants BpSPL3 was shorter,the number of lotus leaves decreased significantly,the number of primary stems increased,the length and number of kernels decreased,and the expression of four-wheeled flower organs was significantly reduced BpSPL3 transgenic strains were found.Real-time fluorescence quantitatively analyzed the expression levels of PNY,LFY,LBD10 and LBD27 genes in the inflorescence of transgenic plants,and the results showed that the expression increased by 2-4 times.It is shown that in the expression of BpSPL3 transgenic Arabidopsis,the PNY,LFY,LBD10 and LBD27 genes may be directly or indirectly involved in flower development and regulated by the BpSPL3 gene.