The Abdomen Transcriptome Sequencing Analysis And The Excavation Of Digestive Function Gene Of Ceracris Kiangsu

Ceracris kiangsu Tsai belongs to the genus Ceracris of Acrididae,is one of the main forest pests,which is extremely harmful to the bamboo of gramineae.At present,chemical control has been mainly undertaken to prevent this species,but due to the im-pact of plant resistance,it has been inadequate.Therefore,it is emergency to explore other more effective prevention and new control methods.Serine protease,as an improtant protease in the insect midgut,mainly including trypsin and chymotrypsin,which have been palyed an important roles in food digestion,innate immune response and involved in many other various physiological and biochemical processes.Because food digestion and innate immune response is crucial to growth and development of insect,a growing number of researchers are interested in insect serine protease.Until now,the research reports of serine protease in orthoptera is mainly in Locusta migratoria,however,the reserach of pests of Ceracris is not involved in.In this paper,based on abdomen transcriptome of Ceracris kiangsu,it is the first time to excavate and analyse a large number of trypsins and chymotrypsins,the CkiaCHY12 gene was cloned by RT-PCR and SMARTer RACE method and carried out various bioinformatics analysis,the conclusions are as follows:1.In abdomen transcriptome of Ceracris kiangsu,the raw reads of each sample of testis and midgut are all greater than 5Gb,finally,we acquire about 290 million clean reads.After assembly,123,927 unigenes are yielded.Of which,the contain of GC is 39.60%,the mean length is 849 bp and the value of N50 is 1,275 bp.Through annotation results of different database,a total of 35,532 unigenes are annotated in UniProt,18,760 unigenes are assigned to 62 clusters of gene ontology(GO),15,125 unigenes are assigned to 25 different clusters of orthologous groups(COG),19,212 unigenes are annotated in KEGG and involved in 288 pathway.Meanwhile,we detect 21,464 potential SSRs.2.The results of differential expression analysis can find:there are 4,765 differential genes between male midgut and testis sample.Of which,2,869 differentially down genes and 1,896 differentially up genes.3.We excavate 5 TRYs and 16 CHYs from transcriptome database.In phylogenetic analysis of TRYs,except LmigTRY3 and CkiaTRY-1,the two kinds of orthoptera insects cluster together;In phylogenetic analysis of CHYs,the two kinds of orthoptera insects are divided into three clades.Tissue expression profile is found,besides CkiaCHYla is highly expressed in hidgut and CkiaTRY1B-is highly expressed in midgut,all of genes are highly expressed in gastric caeca.If it is correct to judge those proteins belong to what kind of protein from substrate specific sites,we can find just CkiaTRY1B,CkiaCHY3a and CkiaTRY1C belongs to trypsins,the rest of other genes belongs to chymotrypsins.4.The length of CkiaCHY12 is 875 bp which include an open reading frame of 831 bp,encoding 276 amino acids.The molecular weight is 28.24KDa,the isoelectric point is 4.33,the atomic number is 3,948.This protein structure is stable,belongs to hydrophobic protein and secreted protein.In the prediction of phosphorylation sites,there are 22 serine(S),5 threonine(T)and 1 tyrosine(Y).And,we have not found N-glycosylation sites.Tissue expression profile analysis indicate that CkiaCHY12 is expressed in all tissues(except testis).Particularly,the expression of CkiaCHY12 in the gastric caeca is 140-fold higher than in the midgut.