Mechanism Of Differences In Phenotype Between Micro-propagated And Conventional Plants Of ‘yanli'strawberry

With the broad application of strawberry?Fragaria×ananassa?micro-propagated plants,people pay more and more attention to the variation of them.The variation of micro-propagated plants of different strawberry varieties was different.‘Yanli'strawberry is a new strawberry variety selected by Shenyang agricultural university which has high quality and disease resistance.In this paper,we investigated the phenotypic differences between micro-propagated and conventional plants in both propagation stage and cultivation stage,and then analyzed the differentially expressed genes in the stem growth points of micro-propagated and conventional plants by using transcriptome sequencing.Using the diploid forest strawberry?F.vesca?as test material,we then analyzed the function of CCD7gene through RNAi technology.Following the results,we preliminary revealed the molecular mechanism that the ability of micro-propagation strawberry was stronger than that of conventional plants.The main results are as follows:1.After observing and investigating the phenotypes of‘Yanli'strawberry micro-propagated plants and conventional plants in both propagation stage and cultivation stage,we concluded that there were many differences between them.In the propagation stage,the number of new stems decreased while the number of stolons increased 3 to 4 times and the number of blades increased by 1.8 times.During the cultivation stage,the first flowering period and fruit ripening period of micro-propagated generation plants were 6 days earlier than that of conventional plants.Besides,the number of leaves increased significantly while the crown diameter decreased significantly.Other growth indexes of plants,height for example,had no significant difference between them,and the fruit yield and quality had no significant difference.2.Using RNA-seq,we analyzed the transcriptomes of stem growth points recorded from micro-propagated original plants and conventional plants.The results showed that compared with conventional plants,1469 genes were up-regulated and 334 genes were down regulated.The log₂value of the expression ratio of Fa CCD7,a key gene for the synthesis of actrolactone,was 1.58,which indicating that the expression level of Fa CCD7 was significantly up-regulated in micro-propagated.3.We cloned FveCCD7 gene from‘Ruegen'of strawberry by RT-PCR,of which the length coding region was 1926 bp,encoding 641 amino acids.The CDS sequence of FveCCD7 gene was identical with that of Fa CCD7 gene spliced by transcriptome sequencing.In addition,we cloned FveCCD8,another key gene for the synthesis of monolactone,from‘Ruegen'of strawberry by RT-PCR.The CDS length of FveCCD8 was 1686 bp,encoding561 amino acids.The sequence of FveCCD8 gene was identical with that of Fa CCD8 gene spliced by transcriptome sequencing.4.The results of qRT-PCR showed that the expressions of FveCCD7 and FveCCD8genes were tissue-specific.The expressions of two gene were high in young organs such as new leaf and new stem while low in old leaf,lowest in root.5.We constructed the RNAi vectors of FveCCD7 and FveCCD8 genes,then via the agrobacterium mediated genetic transformation,obtained one FveCCD7 gene silencing strain and three FveCCD8 gene silencing strains of‘Ruegen'.The phenotypic data showed that both FveCCD7 gene and FveCCD8 gene could inhibit the tillering of strawberry plants.