Comparative Analysis Of Transcriptome On Root Development Of Platycladus Orientalis Seedlings Induced By Root Cutting

The survival rate of plant transplanting is closely related to the development of root system.Root cutting often occurs in the process of seedling transplanting,and is considered to have a positive impact on fruit tree yield and afforestation survival rate.On the other hand,root cutting damages the integrity of seedling root system.Platycladus orientalis is a common tree species used for vegetation restoration.In the process of transplanting,the influence mechanism of root cutting on its survival rate is not clear.Based on the previous research results in the laboratory,this project plans to carry out research in the following three aspects.This study takes 1-year-old Platycladus orientalis Seedlings as the research object,and comprehensively excavates the signal network and key genes of root development induced by root cutting at the transcriptional level by using the new second-generation high-throughput sequencing technology and the third-generation full-length transcriptome sequencing technology;Secondly,according to the differential genes screened by transcriptome sequencing,quantitative primers were designed,and q PCR technology was used to further verify and analyze the gene expression changes of Platycladus orientalis root cutting post-treatment group and control group in different periods(1d,20 d and 60d).Therefore,the temporal and spatial expression law of the gene in root development was determined.Then some key genes were cloned and sequenced to further verify the reliability and accuracy of transcriptome annotation.This study will comprehensively study from different levels of DNA,RNA,protein,physiology and individual,and systematically clarify the molecular regulation mechanism of root development of Platycladus orientalis Seedlings Induced by root cutting.The main results of this study are as follows:(1)Pac Bio SMRT full length transcriptome sequencing,Illumina/Solexa sequencing and identification analysis.Illumina/Solexa sequencing was used to sequence 18 samples(CI in the initial stage of the control group,CLRP in the lateral root generation stage of the control group,CLRE in the lateral root extension stage of the control group;PI in the initial stage of the root cutting group,PLRP in the lateral root generation stage of the root cutting group,PLRE in the lateral root extension stage of the control group),3 biological replicates and 1 mixed sample of pacbiosmrt full-length transcriptome in each treatment.A total of 124643 identical transcripts were obtained,Among them,101688 high-quality full-length transcripts and 22955 low-quality full-length transcripts were obtained after correction of non full-length transcripts.Illumina RNA-seq of 18 samples was completed.According to the non redundant sequence of each sample,56182 transcripts were annotated to 7 databases,and most unigenes were annotated to Nr(97.74%)database.In addition,the Nr database query showed that the sequence with the highest proportion and the closest matching sequence of Platycladus orientalis was North American spruce.All unigenes were annotated.A total of 50 go functional categories included 39532 unigenes,25 cog functional categories included 19763 unigenes,243 KEGG metabolic pathways included16806 unigenes,and 21040 SSRs were found.2446 lncrna transcripts were predicted by CNCI analysis,CPAT analysis,CPC analysis and Pfam protein domain analysis.Using Trans Decoder software to predict the coding region sequence and its corresponding amino acid sequence,there are 47740 complete ORFs.The m RNA characteristics of three lateral root development stages of Platycladus orientalis root cutting treatment group and CK group were revealed for the first time.(2)Screening,expression profiling and functional analysis of differential genes in transcriptomeA total of 4950 differentially expressed genes were found in three development stages of Platycladus orientalis Seedlings after root cutting treatment.According to the gene expression trend,these genes were analyzed by sequence cluster analysis,and they were divided into 8clusters.There were significant differences between root cutting treatment group and CK group at three developmental stages(P<0.05)with fold-change >2,2625 genes were differentially expressed.The sequencing analysis of denovo transcriptome in root tissue at three development stages of root cutting treatment group and CK group showed that the difference analysis between root cutting treatment group and CK group showed that in the initial stage,root cutting treatment caused the start-up expression of a large number of genes.The generation period of lateral roots after root cutting treatment is the most critical period of root development of Platycladus orientalis.The functional enrichment of GO and KEGG of different genes in lateral root development stage in root cutting treatment group showed that they were mainly enriched in cell differentiation and proliferation,carbon metabolism and energy metabolism,cell cycle and plant hormone signal transduction.Genes that may be related to promoting the lateral root growth of Platycladus orientalis seedlings after root cutting(such as GRP-14、PLRR、ACO、SPDs、POD、ZTP-8、AIN and ARE etc.)were found.