Molecular characterization and mating type analysis of oyster mushroom (Pleurotus spp.) using single basidiospores for strain improvement

  • Bhavna Gupta
  • B. P. Niranjan Reddy
  • Anil S. Kotasthane
Original Paper


Existence of variability in morphological traits and growth rate of mycelium of homokaryotic single basidiospores can be exploited for the development of inter-strainal hybrids. We isolated 182 single basidiospores from mushroom bodies of P. sajor-caju, P. florida, P. eous and one wild relative of Pleurotus called Hypsizygusulmaris. The single spores were isolated using a new technique that is less prone to contamination and more efficient than the common techniques used by earlier workers. All the isolates showed a varied range of cultural morphology. Mating types of all the isolates within the species were identified on the basis of hyphal fusion via anastomosis with the tester strains. Four compatible pairs of isolates with well prominent tuft in the contact zone were selected for dikaryon isolation. Dikaryons were used for spawn preparation and mushroom cultivation. The dikaryotic isolates with their replicates were evaluated for spawn run period, yield and biological efficiency. 42 isolates (10 di- and 32 mono-karyotic isolates) were analyzed with RAPD genetic markers. Phenotypic characters and mating types of all the 42 isolates analyzed genetically were correlated with their genetic polymorphism data. The isolates showed very large diversity both at the phenotypic and the genotypic level. Available phenotypic and genotypic data can further help in the selection of monosporous isolates for developing inter-strainal hybrids which can lead to better prospects for genetic improvement in different species of Pleurotus.


Single spore Pleurotus spp. Dikaryon Homokaryotic isolates Random amplified polymorphic DNA (RAPD) 


  1. Aldrich J, Cullis CA (1993) RAPD analysis in flax: optimization of yield and reproducibility using klen Taq 1 DNA polymerase, chelex 100, and gel purification of genomic DNA. Plant Mol Biol Rep 11:128–141CrossRefGoogle Scholar
  2. Bisko NA, Kholodny NG (1989) Selection of Pleurotus ostreatus (Jacq. Fr.) Kumm. In: Abstracts of international symposium on mushroom biotechnology. Nanjing, China, p 21Google Scholar
  3. Brown AJ, Casselton LA (2001) Mating in mushrooms: increasing the chances but prolonging the affair. Trends Genet 17:393–400CrossRefGoogle Scholar
  4. Calvo-Bado L, Noble R, Challen M, Dobrovin-Pennington A, Elliott T (2000) Sexuality and genetic identity in the Agaricus section Arvense. Appl Environ Microbiol 66:728–734CrossRefGoogle Scholar
  5. Calvo-Bado A, Challen MP, Thurston CF, Elliot TJ (2001) RAPD characterization of heterogeneity in spore progenies and sexualities in the genus Agaricus. Mycol Res 105:370–376CrossRefGoogle Scholar
  6. Chiu SW, Chen MJ, Chang ST (1995) Differentiating homothallic Volvariella mushrooms by RFLPs and AP-PCR. Mycol Res 99:333–336CrossRefGoogle Scholar
  7. Chiu SW, Chen MJ, Chang ST (1996) Genetic homogeneity of cultivated strains of shiitake (Lentinula edodes) used in China as revealed by the polymorphism chain reaction. Mycol Res 100:1393–1399CrossRefGoogle Scholar
  8. Fritsche G (1981) Some remarks on the breeding, maintenance of strains and spawn of Agaricus bisporus and Agaricus bitorquis. In: Mushroom Science XI. Proceedings of the eleventh international scientific congress on the cultivation of edible fungi, Australia, pp 367–385Google Scholar
  9. Gharehaghaji AN, Goltapeh EM, Masiha S, Gordan HR (2007) Hybrid production of oyster mushroom Pleurotus ostreatus (Jack; Fries) Kummer. Pakistan J Biol Sci 10:2334–2340CrossRefGoogle Scholar
  10. Ikeda K, Nakamura H, Matsumoto N (2002) Mycelial incompatibility operative in pairings between single basidiospore isolates of Helicobasidium mompa. Mycol Res 107:847–853CrossRefGoogle Scholar
  11. Ikegaya N (1997) Breeding and cultivation of shiitake (Lentinus edodes) mushrooms. Food Res Int 13:335–356CrossRefGoogle Scholar
  12. Ito Y, Fushimi T, Yanagi SO (1998) Discrimination of species and strains of basidiomycete genus Coprinus by Random Amplified Polymorphic DNA (RAPD) analysis. Mycoscience 39:361–365CrossRefGoogle Scholar
  13. Jaindaik CL (1997) History and development of Pleurotus: cultivation in the world and future prospects. In: Upadhyay RC, Singh SK, Rai RD (eds) Current vistas in mushroom biology and production. Mushroom Society of India, Solan, pp 181–191Google Scholar
  14. Kapoor M, Sodhi HS, Dhanda S (1996) Strategies for strain improvement in Pleurotus spp. Mushroom Res 5:57–66Google Scholar
  15. Kavousi HR, Farsi M, Shahriari F (2008) Comparison of random amplified polymorphic DNA markers and morphological characters in identification of homokaryon isolates of white button mushroom (Agaricus bisporus). Pakistan J Biol Sci 11:1771–1778CrossRefGoogle Scholar
  16. Kerrigan RW (1990) Eidence of genetic divergence in two populations of Agaricus bisporus. Mycol Res 94:721–731CrossRefGoogle Scholar
  17. Khush RS, Becker E, Wach M (1992) DNA amplification polymorphisms of the cultivated mushroom Agaricus bisporus. Appl Environ Microbiol 58:2971–2977Google Scholar
  18. Kligman AM (1943) Some cultural and genetic problems in the cultivation of the mushroom Agaricus campestris Fr. Am J Bot 30:745–763CrossRefGoogle Scholar
  19. Kotasthane AS (2003) A simple technique for isolation of Xanthomonas oryzae pv oryzae. J Mycol Plant Pathol 33:277–278Google Scholar
  20. Kotasthane AS, Singh US (2000) Simple technique for single spore isolation and a modified two-point inoculation technique of mating in test tubes. J Mycol Plant Pathol 30:366–369Google Scholar
  21. Kothe E (1996) Tetrapolar fungal mating types: sexes by the thousands. FEMS Microbiol Rev 18:65–87CrossRefGoogle Scholar
  22. Kothe E (2001) Mating-type genes for basidiomycete strain improvement in mushroom farming. Appl Microbiol Biotechnol 56:602–612CrossRefGoogle Scholar
  23. Kronstad JW, Staben C (1997) Mating type in filamentous fungi. Annu Rev Genet 31:245–276CrossRefGoogle Scholar
  24. Kues U, Liu Y (2000) Fruiting body production in basidiomycetes. Appl Microbiol Biotechnol 54:141–152CrossRefGoogle Scholar
  25. Kulkarni RK (1991) DNA polymorphisms in Lentinula edodes, the shiitake mushroom. Appl Environ Microbiol 57:1735–1739Google Scholar
  26. Kumar S (1987) Studies on some mushroom families in the North Western Himalayas. Dissertation, H.P. University, Shimla, IndiaGoogle Scholar
  27. Kumar S, Munjal RL (1981) Studies on evolving new strains of the cultivated mushroom (Agaricus brunnescens Peck) by single spore isolation method. Indian J Mush 7:37–42Google Scholar
  28. Kyung-Ho Ma, Lee Gi-An, Lee Sok-Young et al. (2009) Development and characterization of new microsatellite markers for the oyster mushroom (Pleurotus ostreatus). J Microbiol Biotechnol. doi:10.4014/jmb.0811.604
  29. Loftus MG, Moore D, Elliot TJ (1988) DNA polymorphisms in commercial and wild strains of the cultivated mushroom Agaricus bisporus. Theor Appl Genet 76:712–718CrossRefGoogle Scholar
  30. May B, Royse DJ (1982) Confirmation of crosses between lines of Agaricus brunnescens by isozyme analysis. Exp Mycol 7:283–292CrossRefGoogle Scholar
  31. Nasim G, Malik SH, Bajwa R, M Afzal, Mian CG (2001) Effect of three different culture media on mycelia growth of oyster Chinese mushroom. J Biol Sci 1:1130–1133CrossRefGoogle Scholar
  32. Pahil VS (1997) Strain improvement in tropical Agaricus: techniques selection of genetically diverse parents for hybridization in Agaricus bisporous. Mushroom Res 12:19–26Google Scholar
  33. Perisamy K, Nataranjan K (2003) Strain improvement of Pleurotus djamor (Fr.) boedijn var. roseus corner using conventional breeding technique. In: Upadhyay RC, Singh SK, Rai RD (eds) Current vistas in mushroom biology and production. Mushroom Society of India, Solan, pp 35–46Google Scholar
  34. Shahid MN, Abbasi NA, Saleem N (2006) Effect of different methods of compost preparation and lime concentration on the yield of Pleurotus sajor-caju. Int J Agr Biol 8:129–131Google Scholar
  35. Sigoillot C, Camarero S, Vidal T, Record E, Asther M, Perez-Boada M et al (2005) Comparison of different fungal enzymes for bleaching high-quality paper pulps. J Biotechnol 115:333–343CrossRefGoogle Scholar
  36. Singh SK, Doshi A, Yadav MC, Kamal S (2006) Molecular characterization of speciality mushroom of Western Rajasthan, India. Curr Sci 91:1225–1230Google Scholar
  37. Sonnenberg ASM (2000) Genetics and breeding of Agaricus bisporus. Mush Sci 15:25–39Google Scholar
  38. Wang SS, Anderson NA (1972) A Genetic analysis of sporocarp production in Pleurotus sotreatus. Mycologia 64:521–528CrossRefGoogle Scholar
  39. Yadav M, Tripathi YK, Verma RN, Upadhyay RC, Dhar BL (2003) Molecular breeding for development of genetically improved strain and hybrids of Agaricus bisporous. In: Upadhyay RC, Singh SK, Rai RD (eds) Current vistas in mushroom biology and production. Mushroom Society of India, Solan, pp 261–274Google Scholar

Copyright information

© Springer Science+Business Media B.V. 2010

Authors and Affiliations

  • Bhavna Gupta
    • 1
  • B. P. Niranjan Reddy
    • 2
  • Anil S. Kotasthane
    • 3
  1. 1.Evolutionary Genomics and Bioinformatics Laboratory, Division of Genomics and BioinformaticsNational Institute of Malaria Research (NIMR)New DelhiIndia
  2. 2.Vector Control DivisionNational Institute of Malaria Research (NIMR)New DelhiIndia
  3. 3.Department of BiotechnologyIndira Gandhi Krishi VishwavidyalayaRaipur (C.G.)India

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