Mapping Of Mitochondrial Genome, Transcriptome And Mitochondrial Transcriptome Of Teleogryllus Emma

Teleogryllus emma (Ohmachiet Matsumura1951) belongs to Grylloidea, Orthoptera, can be ultilized as medical, edible and animal foods resources. It is widely distributed in China. With a series of advantages, such as large body size, easy dissection, non-diapause, T. emma has been used extensively in the study of insects’ physiological functions, embryo development, endocrine, toxicology and so on. Ye Wei et al. determined and analysed the mtDNA of T. emma in2008. However, the research and analysis about transcriptome and mitochondrial transcript mapping have not been reported.In this paper, we carried out the research on molecular studies of three major areas:mtDNA, transcriptome and mitochondrial transcript mapping. The main conclusions of the study are as follows:(1) The determined mitochondrial genome sequences of T. emma is15,697bp, encoding a total of37genes (including13protein-coding genes and22tRNAs) and a length of954bp non-coding regions. The base compositon shows AT-bias; the13PCGs use TTA as start codon except CO I, and typical TAN as stop codon; NNU and NNA are used as protein codon frequently; the most widely used amino acid of PCGs is Leu (14.69%); all the tRNA could form typical clover structure except tRNASer(AGN). The T. emma annotation results are generally similar with T. emma of Ye Wei in base compositions, AT content, protein coding gene, tRNA secondary structures and so on.(2) The gene rearangement of T. emma is the third type in Orthoptera mitochondrial genome that has been found so far which is consistant with Ye Wei’s research (the inversion among tRNAGlu、tRNASer(AGN) and tRNAAsn), which is quite different from the traditional ARNSEF arrangement of most Caelifera species. tRNAGlu、tRNASer(AGN) and tRNAAsn are encoded by N strand, and other species of Orthoptera encoded by J strand. That may be the common characteristics of Grylloidea mitochondrial genome.(3) The AT-rich region of T. emma is954bp in length with a22bp repeat units which repeats2.5times located in15,300-15,355bp. After compared with the AT-rich region of Schistucerca, we identified4conversed sequence structures in T. emma. Stem loop structure has been found in the AT-rich regio of T. emma. The stem composed of15bp, the ring composed of18bp.(4) By sequencing of male and female adults transcriptome, thousands of clean reads have been found, including52,507,034pair reads in female adult,51,524,978pair reads in male adult. After assembling these data, we obtained64,806in female adult,63,760in male adult and merged62,854All-Unigene.(5)30,180All-Unigene have been annotated accouting for45%throngh blasting against various database, including Nr, SwissProt, KEGG and COG. Most of the Unigene sequences were matched with the Nr database, with a total of26,386;10,576were matched with COG;19,776were matched with KEGG metabolic pathways;13,560were matched with GO.(6) For the SSR analysis, we find out a total of4,803SSR loci which length distributed between11-24bp. The number of each nucleotide were as follows:835Mono-nucleotide,1,301Di-nucleotide,2,258Tri-nucleotide,131Quad-nucleotide,165Penta-nucleotide,113Hexa-nucleotide. The most frequently used categories of repeat units are CCG/CGG, A/T, AC/GT, AG/CT.(7) For the detection of All-Unigene SNP, female adult has43,047variations sites,26,824base transitions and16,223transversions; male adult has40,761variation sites,25,282base transition and15,479transversions. The number and type distribution of female and male adults of T. emma remain the same. The number of transitions (>60%) is greater than transversions. A-T transversion mutation has higher proportion than others:female adult is11.36%while male adult is11.23%.(8) A total of16,326differentially expressed genes was found,9,698were up-regulated in female,6,528were up-regulated in male.4,319DEGs were assigned to GO classification,18,823involved in biological process,9,382annotated into molecular function, and5,635assigned to cellular. In molecular function, most unigene were involved in binding function. Differential gene in KEGG metabolism pathway shows6,455All-Unigene annotated into256different metabolism patheways in which46different pathways enriched, including909differential genes in metabolism pathway at most. Gene CYP314A1and CYP18A1which belong to the CYP450enzyme system were up-regulated in female adult, suggesting that it may be involved in the biosynthesis of the insect sex hormones. Gene JHAMT, EO and Nvd which were up-regulated in male adult, suggesting it may be involved in the biosynthesis of the juvenile hormone, molting hormone and cholesterol.(9) Through mitochondrial transcript mapping we found that15,305of15,697sites with coveraged at about97.5%, tRNAIle、tRNAGln and part of tRNAMet, AT-rich region are not covered, indicating that the transcription efficiency is higher than in protein-coding genes and rRNA plays an important role in ribosome assemble. Signficant differences in16S rRNA coverage may be related to its secondary structures. The two pairs of overlapping genes, ATP8/ATP6, ND4/ND4L are transcribed from a bicistronic mRNA respectively. T. emma has five large transcriptional units, they can be processed into mature transcripts through tRNA punctuation model.This research sequenced and analysed the mtDNA and finally validates the tRNA rearrangement in T. emma. A large number of Unigene obtained from transcriptomes of female and male adults, and the function of differently expressed genes analyzed and obtained the genes related to growth, development and sexual differiation. The mitochondrial transcript mapping of T. emma, overcoming the shortage of mitochondrial genome annotation.