Analysis Of The Full-length Transcriptome And Mitochondrial Transcriptome Of Three Orthopteran Insects

Locusts are one of the most destructive agricultural pestswith worldwide distribution,and it also have high potentiality in nutrition and econommics.Using Pacbio RS II platform,full-length transcriptome of Oxya chinensis,Acrida cinereaand Atractomorpha sinensiswere sequenced.In addition,the high quailty full-length transcriptswere mappedto mitochondrial genomes in three locusts,respectively,and we built a straightfoward to investigate the mechanism on RNA processing of mitochondrial gene.The main results in this study are as follows:1)Based on Pacbio RS II platform,a total of 13.06 Gb,12.57 Gb and 12.61 Gb of clean reads;365,726,360,844 and 387,390 reads of insert(ROI);153,161,137,361 and 137,361 of full-length non-chimeric reads(FLNC)were obtainedin O.chinensis,A.cinereaand A.sinensis,respectively.After transcripts clusterand error correction,a total of 42,589,38,404 and 46,365 high-quailty full-length transcripts were generated,with the N50 length of3,372bp,3,527bp and 3,367 bp in O.chinensis,A.cinereaand A.sinensis,respectively.Moreover,80.0%,82.1%and 86.7%of the BUSCO Arthropoda gene annotations wereidentified.Based on these findings,we generated an ideal integrity transcripts which could be used for downstream analysis like functional annotation.2)In O.chinensis,A.cinereaand A.sinensis,6,612,5,499 and 5,318 IncRNAs were predicted,with the N50 length of 3,379bp,3,452 bp and 3,357 bp.Only 12,10 and 8 IncRNAs were highly homologous with D.Melanogaster in NONCODE database,respectively.The expression profiles of different development in locusts were analyzed,and manydifferentially expressed InRNAs were detected.These findings show that full-length transcriptome sequencing can efficiently capture more and longer candidate non-coding transcripts,and these IncRNAs could be used as a reference database for further studies in other related locusts species.3)Using TransDecoder,35,995,32,817 and41,125open reading frames(ORFs)were identified,which included 24,955,23,161 and 28,281 complete ORFs in O.chinensis,A.cinereaand A.sinnsis,respectively.High-quailty full-length transcripts were then mapped to six databses to get the annotation information of the transcript,a total of 34,528(81.07%),32,207(83,86%)and3 9,326(84.82%)transcripts weresuccessfully annotated,which could provide a useful perspective elucidation on physiological metabolism,gene regulation,phyletic evolution and molecular marker development.In addition,based on these annotations,many putative full-length transcripts related to locusts' immune,growth,development and nutritional components were identified,which could provided a molecular foundationto develop new targets for the biological control of locusts,and the reports of genomic information forprospective commercial locusts seem timely(currentlylacking)and helpful to understand the potential applications ofgrasshoppers as feed,food or medicine.4)In order to obtain more comprehensive and effective transcripts of the three locusts at different developmental stages,based on high-quailty full-length transcripts,we combined the clean reads from RNA-seq transcriptomes to realize the quantification of the three locust transcripts at different stages of development.Between nymph and adult stage,the full-length transcriptsrelated to growth and development were exhibited higher expression levels at the nymphstage,and the transcripts associated withenvironmental adaptability,metabolise and iron ion transportshowed increased transcription inadults.Between female male adults,the full-length transcriptsrelated to metabolise were exhibited higher expression levels at the female adults,and the transcripts associated withenvironmental adaptability and adult chitin-based cuticle in A.cinereaand A.sinensis,respectively?5)The high-quailty full-length transcripts were mapped to mitochondrial genomes in three locusts,respectively.We constructed the first full-length transcription map of Orthoptera insects mitochondria,and the structure of the mapped full-length transcript were analyzed.Most of the results were not consistent with the previous studies,while to the best of our knowledge,some findings were reported for thefirst time in this study.Four kinds of primary transcripts exist in O.chinensis and A.cinerea' mitochondrial,and six kind of primary transcripts exist in A.sinensis' mitochondrial.The bicistronic of nd4L/nd4,atp6/atp8?nd6/cytB,and the tricistronic atp8-atp6-cox3 transcripts are likely to function as intermediate product of transcripts instead of mature transcripts in RNA processing.Moreover,this study speculates that tRNAs preceding mRNAs or rRNAs inthe polycistronic transcripts are removed both by5' to 3' and 3' to 5'cleavage,and the overall processing direction of polycistronic transcripts both have5' to 3' and 3' to 5'terminals.These findings were challenged to fundamental concepts of mitochondrial gene transcription and RNA processing.