Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Cordyceps militaris (Hypocreales: Cordycipitaceae): transcriptional analysis and molecular characterization of cox1 and group I intron with putative LAGLIDADG endonuclease

  • 355 Accesses

  • 4 Citations


The full-length cytochrome c oxidase subunit I gene (cox1) containing a group I intron was isolated from an important medical fungus Cordyceps militaris (Cordycipitaceae). The open reading frame (ORF) of 1,593 nucleotides encoded a predicted protein COX1 of 530 amino acids. The group I intron encoded a putative homing endonuclease (HE) with two LAGLIDADG motifs. RT-PCR and Northern analysis showed a mature transcript of spliced cox1. Both 5′exon-intron and intron-3′exon junctions were also found by RT-PCR, suggesting the possible presence of unspliced cox1 RNA in total RNA. Sequence comparison by BLASTn showed that the coding region of cox1 (CRcox1) of C. militaris had significant similarities to those of related species (such as Cordyceps bassiana and C. brongniartii), while the intron had no significant homologous sequences of Cordycipitaceae fungi in NCBI database. The phylogenetic tree based on the CRcox1 confirmed the present taxonomic status of related species, but the cox1 introns were phylogenetically distinct. Compared to C. bassiana and C. brongniartii, the cox1 intron of C. militaris exhibited specific splicing site and different intronic ORF. The analysis of the folding RNA structures of the known cox1 introns from Cordyceps species showed different base pairs and conserved regions (P1–P10) in their structures. The present results provide useful information on the studies of cox1 intron splicing and Cordyceps evolution.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4


  1. Adams KL, Clements MJ, Vaughn JC (1998) The Peperomia mitochondrial coxI group I intron: timing of horizontal transfer and subsequent evolution of the intron. J Mol Evol 46:689–696

  2. Akashi H, Matsumoto S, Taira K (2005) Gene discovery by ribozyme and siRNA libraries. Nat Rev Mol Cell Biol 6:413–422

  3. Beagley CT, Okada NA, Wolstenholme DR (1996) Two mitochondrial group I introns in a metazoan, the sea anemone Metridium senile: one intron contains genes for subunits 1 and 3 of NADH dehydrogenase. Proc Natl Acad Sci USA 93:5619–5623

  4. Brzezinski P, Gennis RB (2008) Cytochrome C oxidase: exciting progress and remaining mysteries. J Bioenerg Biomembr 40:521–531

  5. Cech TR, Damberger SH, Gutell RR (1994) Representation of the secondary and tertiary structure of group I introns. Nat Struct Biol 1:273–280

  6. Chen Y, Hu B, Xu F, Zhang W, Zhou H, Qu L (2004) Genetic variation of Cordyceps sinensis, a fruit-body-producing entomopathogenic species from different geographical regions in China. FEMS Microbiol Lett 230:153–158

  7. Chevalier B, Stoddard B (2001) Homing endonucleases: structural and functional insight into the catalysts of intron/intein mobility. Nucleic Acids Res 29:3757–3774

  8. Chevalier B, Monnat R, Stoddard B (2005) The LAGLIDADG homing endonuclease family. Nucleic Acid Mol Biol 16:33–47

  9. Felsenstein J (1981) Evolutionary trees from DNA sequences: a maximum likelihood approach. J Mol Evol 17:368–376

  10. Fukami H, Chen CA, Chiou C, Knowlton N (2007) Novel group I introns encoding a putative homing endonuclease in the mitochondrial cox1 gene of Scleractinian Corals. J Mol Evol 64:591–600

  11. Ghikas DV, Kouvelis VN, Typas MA (2006) The complete mitochondrial genome of the entomopathogenic fungus Metarhizium anisopliae var. anisopliae: gene order and trn gene clusters reveal a common evolutionary course for all Sordariomycetes, while intergenic regions show variation. Arch Microbiol 185:393–401

  12. Ghikas DV, Kouvelis VN, Typas MA (2010) Phylogenetic and biogeographic implications inferred by mitochondrial intergenic region analyses and ITS1–5.8S-ITS2 of the entomopathogenic fungi Beauveria bassiana and B. brongniartii. BMC Microbiol 10:174–188

  13. Golden BL, Gooding AR, Podell ER, Cech TR (1998) A preorganized active site in the crystal structure of the Tetrahymena ribozyme. Science 282:259–264

  14. Gong ZH, Su Y, Huang L (2009) Cloning and analysis of glyceraldehyde-3-phosphate dehydrogenase gene from Cordyceps militaris. Afr J Agric Res 4:402–408

  15. Haugen P, Simon DM, Bhattacharya D (2005) The natural history of group I introns. Trends Genet 21:111–119

  16. Joyce GF (2002) The antiquity of RNA-based evolution. Nature 418:214–221

  17. Kuo HC, Su YL, Yang HL, Chen TY (2005) Identification of Chinese medicinal fungus Cordyceps sinensis by PCR-single-stranded conformation polymorphism and phylogenetic relationship. J Agric Food Chem 53:3963–3968

  18. Li ZJ, Zhang Y (2005) Predicting the secondary structures and tertiary interactions of 211 group I introns in IE subgroup. Nucleic Acids Res 33:2118–2128

  19. Li L, Pischetsrieder M, St Leger RJ, Wang C (2008) Associated links among mtDNA glycation, oxidative stress and colony sectorization in Metarhizium anisopliae. Fungal Genet Biol 45:1300–1306

  20. Lunt DH, Zhang DX, Szymura JM, Hewltt GM (1996) The insect cytochrome oxidase I gene: evolutionary patterns and conserved primers for phylogenetic studies. Insect Mol Biol 5:153–165

  21. Michel F, Westhof E (1990) Modelling of the three-dimensional architecture of group I catalytic introns based on comparative sequence analysis. J Mol Evol 216:585–610

  22. Nesbø CL, Doolittle WF (2003) Active self-splicing group I introns in 23S rRNA genes of hyperthermophilic bacteria, derived from introns in eukaryotic organelles. Proc Natl Acad Sci USA 100:10806–10811

  23. Nikoh N, Fukatsu T (2001) Evolutionary dynamics of multiple group I introns in nuclear ribosomal RNA genes of endoparasitic fungi of the genus Cordyceps. Mol Biol Evol 18(9):1631–1642

  24. Nishida H, Sugiyama J (1995) A common group I intron between a plant parasitic fungus and its host. Mol Biol Evol 12:883–886

  25. Ogawa S, Matsuo K, Angata K, Yanagisawa K, Tanaka Y (1997) Group-I introns in the cytochrome c oxidase genes of Dictyostelium discoideum: two related ORFs in one loop of a group-I intron, a cox1/2 hybrid gene and an unusually large cox3 gene. Curr Genet 31:80–88

  26. Posada D, Crandall KA (1998) Modeltest: testing the model of DNA substitution. Bioinformatics 14:817–818

  27. Rot C, Goldfarb I, Ilan M, Huchon D (2006) Putative cross-kingdom horizontal gene transfer in sponge (Porifera) mitochondria. BMC Evol Biol 6:71

  28. Saldanha R, Mohr G, Belfort M, Lambowitz AM (1993) Group I and group II introns. FASEB J 7:15–24

  29. Saraste M (1990) Structural features of cytochrome oxidase. Q Rev Biophys 23:331–366

  30. Stensrud Ř, Hywel-Jones NL, Schumacher T (2005) Towards a phylogenetic classification of Cordyceps: ITS nrDNA sequence data confirm divergent lineages and paraphyly. Mycol Res 109:41–56

  31. Sung GH, Hywel-Jones NL, Sung JM, Luangsa-Ard JJ, Shrestha B, Spatafora JW (2007) Phylogenetic classification of Cordyceps and the Clavicipitaceous fungi. Stud Mycol 57:5–59

  32. Ujita M, Katsuno Y, Suzuki K, Sugiyama K, Takeda E, Yokoyama E, Hara A (2006) Molecular cloning and sequence analysis of the b-1, 3-glucan synthase catalytic subunit gene from a medicinal fungus, Cordyceps militaris. Mycoscience 47:98–105

  33. Vicens Q, Cech TR (2006) Atomic level architecture of group I introns revealed. Trends Biochem Sci 31:41–51

  34. Vicens Q, Paukstelis PJ, Westhof E, Lambowitz AM, Cech TR (2008) Toward predicting self-splicing and protein-facilitated splicing of group I introns. RNA 14:2013–2029

  35. Wang Z, He Z, Shen Q, Gu Y, Li S, Yuan Q (2005) Purification and partial characterization of recombinant Cu, Zn containing superoxide dismutase of Cordyceps militaris in E. coli. J Chromatogr B 826:114–121

  36. Watanabe KI, Ehara M, Inagaki Y, Ohama T (1998) Distinctive origins of group I introns found in the COX1 genes of three green algae. Gene 213:1–7

  37. Woo PC, Zhen H, Cai JJ, Yu J, Lau SK, Wang J, Teng JL, Wong SS, Tse RH, Chen R, Yang H, Liu B, Yuen KY (2003) The mitochondrial genome of the thermal dimorphic fungus Penicillium marneffei is more closely related to those of molds than yeasts. FEBS Lett 555:469–477

  38. Zhang YJ, Xu LL, Zhang S, Liu XZ, An ZQ, Wang M, Guo YL (2009) Genetic diversity of Ophiocordyceps sinensis, a medicinal fungus endemic to the Tibetan Plateau: implications for its evolution and conservation. BMC Evol Biol 9:290

  39. Zheng ZL, Huang CH, Cao L, Xie CH, Han RC (2011) Agrobacterium tumefaciens-mediated transformation as a tool for insertional mutagenesis in medicinal fungus Cordyceps militaris. Fungal Biol 115:265–274

  40. Zhou XW, Gong ZH, Su Y, Lin J, Tang KX (2009) Cordyceps fungi: natural products, pharmacological functions and developmental products. J Pharm Pharmacol 61:279–291

Download references


This work was supported by the Research Project of Guangdong Province (Grant 2008B040200017), and by the Guangdong Province-Chinese Academy of Sciences Comprehensive Strategic Cooperation Project (Grant 2009B091300015).

Author information

Correspondence to Richou Han.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Zheng, Z., Jiang, K., Huang, C. et al. Cordyceps militaris (Hypocreales: Cordycipitaceae): transcriptional analysis and molecular characterization of cox1 and group I intron with putative LAGLIDADG endonuclease. World J Microbiol Biotechnol 28, 371–380 (2012). https://doi.org/10.1007/s11274-011-0829-9

Download citation


  • Cordyceps militaris
  • cox1
  • Group I intron
  • Splicing
  • LAGLIDADG endonuclease