Abstract
Identification of monokaryons and their mating types and discrimination of hybrid offspring are key steps for the crossbreeding of Pleurotus tuoliensis (Bailinggu). However, conventional crossbreeding methods are troublesome and time consuming. Using RNA-seq technology, we developed new expressed sequence tag-simple sequence repeat (EST-SSR) markers for Bailinggu to easily and rapidly identify monokaryons and their mating types, genetic diversity and hybrid offspring. We identified 1110 potential EST-based SSR loci from a newly-sequenced Bailinggu transcriptome and then randomly selected 100 EST-SSRs for further validation. Results showed that 39, 43 and 34 novel EST-SSR markers successfully identified monokaryons from their parent dikaryons, differentiated two different mating types and discriminated F1 and F2 hybrid offspring, respectively. Furthermore, a total of 86 alleles were detected in 37 monokaryons using 18 highly informative EST-SSRs. The observed number of alleles per locus ranged from three to seven. Cluster analysis revealed that these monokaryons have a relatively high level of genetic diversity. Transfer rates of the EST-SSRs in the monokaryons of closely-related species Pleurotus eryngii var. ferulae and Pleurotus ostreatus were 72% and 64%, respectively. Therefore, our study provides new SSR markers and an efficient method to enhance the crossbreeding of Bailinggu and closely-related species.
Highlights
Pleurotus tuoliensis [1], known as Bailinggu, is one of the many significant, choice edible mushrooms [2]
We found that culturing liquid mycelia for 11 days was more effective than the traditional seven days
The rate of the protoplast-derived monokaryon development from regenerated colonies was more than 93% using 11-day-old liquid mycelia, compared to about 60%
Summary
Pleurotus tuoliensis [1], known as Bailinggu, is one of the many significant, choice edible mushrooms [2]. Bailinggu was first cultivated successfully in China in 1986 [3]. More research has been devoted to its genetic diversity, breeding and cultivation [1,2,3,4,5]. Only six varieties are used for commercial cultivation. Strengthening the genetics and breeding of Bailinggu is urgently needed. Crossbreeding plays an important role in mushroom breeding [6,7], but it is seldom done with
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