Cloning And Expressiong Of Genes Related To Stone Cells And Bioinformatic Analysis Of HCT Gene Family

Stone cells is one of the important indicators that impact the quality of pear.The content of lignin which is the main component of stone cells is affected by genetic characteristics,cultivation practices and environment.In this study,we analysis stone cells releated genes by bioinformatics methods and fand the key genes associated with the formation of stone cells.Meanwhile,we had cloned the PbMYB85 gene in 'Dangshansuli'and constructed the expression vectors Pcamibia1301-PbMYB85.This vector was then used in the tobacco genetic transformation to explore the PbMYB85 gene function.The main findings were as follows:1 A total of 82 HCT genes were identified in pear,most of which have one or two exons,and all with the conserved domains HXXXD and DFGWG.Based on the structural characteristics and phylogenetic analysis of these sequences,the HCT gene family genes could be classified into four main groups.Structural analysis also revealed that 25%of HCT genes share a MYB binding site.Expansion of the HCT gene family mostly occurred before the divergence between Arabidopsis and Rosaceae,with WGD or segmental duplications events playing the most important role in the expansion of the HCT gene family in pear.Six of the 82 HCT genes found were verified by qRT-PCR to correspond to the pattern of stone cell formation during pear fruit development.2 According to QTL interval mapping and expression analysis,we found that MYB transcription factor Pbr041889.1 may be incolved in the regulation of lignin in pear stone cells,we named it as PbMYB85.It contained the typical characteristics of MYB transcription factor,and belong to MYB transcription factor family.PbMYB85 gene showed similar expression in multiple variety of pears,and increased expression in the early stages and than decreased gradually.The exppression of MYB is positively correleated with the content of lignin and stone cells in pear.So we speculate that it may be involved in the formation of stone cells in pear fruit.3 This study successfully cloned PbMYB85 gene and constructed the expression vector pCAMBIA301-PbMYB85,and this vector had trasformed in to the tobacco leaves via Agrobacterium-mediated genetic transformation method and several independent transgenic lines were obtained,which is significantly valuable to explore the gene function of PbMYB85 in depth.