Study On The Technical System Of Industrialized Seedling Raising And The Genetic Variation Of Tube Seedlings In Successive Generations Of Atractylodes Lancea(Thunb.)DC

Atractylodes lancea(Thunb.)DC is a perennial plant of Atractylodes in Compositae.In recent years,the wild resources of A.lancea are on the verge of extinction.The lack of seedling source is one of the main reasons that restrict the progress of artificial cultivation.In this paper,we used A.lancea as experimental material,optimized the plant-based seedling technology system,analyzed the genetic stability and DNA methylation changes in the process of subculture of test tube seedlings of A.lancea,and explored the genetic variation mechanism in the process of subculture of A.lancea from the molecular level.The main results are as follows:1.Study on the optimization of the technical system of the factory seedling of Atractylodes lancea(Thunb.)DC.The results showed that the best proliferation medium was MS+1.0 mg/L6-BA+0.1 mg/LNAA,and the best rooting medium was 1/2MS+0.5 mg/LNAA+1.0 g/L activated carbon;compared with the traditional tissue culture,sugar free tissue culture can effectively improve the photosynthetic autotrophic ability of the seedlings of A.lancea.When using 1/2MS as the culture medium for sugar free tissue culture,the plant growth and root system development level are the best;when the ratio of the matrix of the hole plate is V(vermiculite):V(coconut bran):V(peat soil)=1:1:1,the survival rate of A.lancea is the highest,which is 92.10%.2.Genetic stability analysis of the subculture of test tube seedlings of Atractylodes lancea(Thunb.)DC.It was found that the genetic diversity of 10 samples with different subculture times was low and had high genetic stability.Among them,the genetic similarity coefficient between the first generation and the tenth generation samples is the lowest,and the genetic relationship is far away;but from the fourth generation,the bands of different primer amplification patterns change at individual sites,and the change is more obvious with the increase of the number of generations.After long-term subculture,the methylation level of test tube plantlet of A.lancea decreased,and the demethylation mode(18.87%)and methylation mode(13.46%)coexisted,but the demethylation mode was the main mode,which indicated that 18.87%of genes were reactivated and expressed,and 13.46%of genes were inhibited after long-term subculture.3.Transcriptome analysis of subculture of test tube plantlet of Atractylodes lancea(Thunb.)DC.The results showed that 80 285 unigenes and 58 951 differentially expressed genes were obtained by transcriptome sequencing,of which 36 859 were up regulated and down regulated 22 092;there are 43 351 differentially expressed genes annotated into the biological process,cell components and molecular functions of go library,which are closely related to binding,catalytic activity,metabolic process,cell process and biological regulation;pathway analysis shows that 15 629 differentially expressed genes are involved in MAPK signal pathway,biosynthesis and antibiosis of plant hormone signal transduction secondary metabolites There were significant differences in the biosynthesis of phytohormones and the metabolism of microorganisms in different environments.Due to the influence of exogenous hormones,sucrose,agar,pH value and other factors,the long-term subcultured test tube seedlings of A.lancea showed abnormal expression in secondary metabolites,antibiotic biosynthesis,MAPK signal pathway,neurotrophin signal pathway and plant hormone signal pathway.