A Study On The Flammability Characteristics Of Artemisia Argyi By RNA Sequencing

Objective: To analyze Artemisia argyi(A.argyi)transcriptome data and to search for synthetic enzyme genes that control combustible substances by RNA-seq technology,which in order to explore the transcriptomic basis of the flammability characteristics of A.argyi,and to provide the experimental basis for the molecular biology of leaves as raw material in moxibustion.Methods: After total RNA extracted from fresh leaves,roots and stems of A.argyi,c DNA libraries of the three tissues were established for high-throughput sequencing on Illumina Hi Seq 4000 platform.Clean reads were filtered by raw reads obtained from sequencing,and clean reads was assembled and removed redundancies by Trinity and Tgicl softwares,termed Unigene.To acquire unigene functional annotations,unigenes were aligned to protein databases,including Nt、Nr、COG、KEGG and Swiss-Prot using Blastx.The gene encoding enzymes involved in terpenoids,cellulose and lignin biosynthesis were identified and analyzed according to the Ko number of KEGG pathway annotation.The genes which encode key enzymes involved in terpenoids biosynthesis were cloned by RT-PCR for verifying the correctness of transcriptome data.Results: Illumina high-throughput sequencing of the A.argyi transcriptome generated approximately 33.46 Gb clean reads.A total of 67,446 annotated unigenes were acquired significant hits in seven different databases,consist of 20,091(20.13%)by COG,25,638(25.69%)by GO and 44,750(44.84%)by KEGG.With KEGG pathway annotation,116 unigenes encoding AATC、HMGS、HMGR、PMK、MVD、DXS、DXR、CMS、CMK、MCS、HDS、HDR、IDI、GPPS and FPPS enzymes,were mapped to terpenoid backbone biosynthesis;119 unigenes encoding Su Sy、Ces A、GPI、PGM and UGP2 enzymes,were mapped to the synthesis of cellulose and 256 unigenes encoding PAL、C4H、4CL、HCT、C3H、CCo AOMT、COMT、F5H、CCR and CAD enzymes,were mapped to the synthesis of lignin.In terpenoid biosynthesis,the genes encoding PMK、MVD、DXR、CMS、CMK and HDS were higher in the leaves than in the stems and roots.We successfully cloned HMGR、MVD、DXS、DXR、HDS and HDR genes from A.argyi leaves for verifying the reliability of transcriptome.Conclusion: 1.Many of the key enzyme genes involved in invovled in terpenoids biosynthesis,cellulose and lignin are contained in the wormwood,which is the molecular biology basis of the flammability characteristics of A.argyi.2.The key enzyme genes invovled in terpenoids biosynthesis,such as PMK,MVD,DXR,CMS,CMK,and HDS,which control the synthesis of volatile oil from A.argyi,are higher in the leaves than in the roots and stems of A.argyi.This may be the scientific basis for selecting A.argyi lesf as raw materials in moxibustion.3.The genes invovled in terpenoids,cellulose and lignin biosynthesis in roots and stems illustrate that roots and stems have flammability properties,prompting root and stem have development potential for raw materials in moxibustion.