The Analysis,Cloning And Expression Of The Functional Genes From Hirsutella Sinensis

Cordyceps sinensis is widely used as the Chinese traditional medicine in the treatment of certain diseases.Due to rarity and consequent high cost of the natural C.sinensis,many scientists have extensively examined its life cycle with the aim of developing techniques for isolating fermentable strains.Numerous researches indicated that Hirsutella sinensis is the anamorph of C.sinensis.The products from cultured H.sinensis have shown similar pharmacological efficacy comparable to that of natural C.sinensis.In order to fully elucidate the functional complexity of the H.sinensis,the transcriptome of the mycelium of H.sinensis L0106 in different cultivation phase(3D,6D and 9D)was sequenced using next-generation sequencing technology RNA_seq,a powerful tool for transcriptome analysis,respectively.We totally obtained 70,743,026 reads,in which 26,577,780 reads with an average length of 90 nt from the 3D transcriptome,27,355,558 reads with an average length of 90 nt from the 6D transcriptome and 16,809,688 reads with an average length of 75 nt from the 9D transcriptome.In the transcriptome de novo analysis,20822 Unigenes(All)were obtained with a mean size of 1013 nt and annotated.At last,12980 Unigenes(All)were annotated with the protein annotation,6627 with COG functional annotation,1353 with GO functional annotation and 8724 with KEGG Pathway annotation.In the transcriptome reference genome analysis,clean reads from different transcriptome were mapped to reference genome and gene sequences(the genome of the H.sinensis L0106 our laboratory had sequenced)respectively.The gene expression profiles of H.sinensis during different developmental stages were compared with a digital gene expression system.The assembled,annotated transcriptome sequences and gene expression profiles provide an invaluable resource for the identification of H.sinensis genes involved in its medical function.Furthermore,the anabolism pathways of mannitol,cordycepin and purine nucleosides in H.sinensis were predicted and mapped.In total,39 functional genes from the pathways were amplified and expressed in the E.coli BL21(DE3).