Effects of Medium Composition and Genetic Background on Agrobacterium-Mediated Transformation Efficiency of Lentinula edodes.

Lianlian Yan,Haiyang Liu,Yan Zhou,Ruiping Xu,Yuhua Gong,Yinbing Bian,Shenghong Dai

doi:10.3390/genes10060467

Lianlian Yan, Haiyang Liu + Show 5 more

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https://doi.org/10.3390/genes10060467

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Journal: Genes	Publication Date: Jun 19, 2019
Citations: 14	License type: CC BY 4.0

Affiliation: Huazhong Agricultural University

Abstract

The establishment of genetic transformation method is crucial for the functional genomics research in filamentous fungi. Although the transformation method has been developed in several types of fungi, a highly efficient and convenient transformation system is desperately needed in Lentinula edodes. Present work established the Agrobacterium-mediated transformation (ATMT) of basidiomycete L. edodes in both monokaryon and dikaryon mycelia by using constructed binary plasmid pCAMBIA-1300-GFP. Then, the transformation efficiency of ATMT was evaluated by using different mediums for recipient incubation and different varieties of L. edodes. The results showed that in dikaryon strain W1, the positive hygromycin-resistant transformants was observed in all medium with the positive frequency of selected transformants that ranged from 0 to 30%. While in the monokaryon strain W1-26, only the millet medium group obtained positive transformants with a positive frequency of 75.48%. Moreover, three dikaryotic wild strains (YS55, YS3334, and YS3357) and two dikaryotic cultivated strains (W1 and S606) showed the highest transformation efficiency, with 32.96% of the germination frequency, and 85.12% of positive frequency for hygromycin-resistant transformants. This work demonstrated that Agrobacterium-mediated transformation was successfully performed in L. edodes, and the genotype of recipients as well as the medium for mycelial incubation were suggested to play key roles in determining the transformation efficiency. These findings may provide new avenues for the genetic modification of edible mushroom and may extend the cognition of DNA-mediated transformation in filamentous fungi.

Highlights

Filamentous fungi encompass diverse groups and are widely distributed in nature
In the second selection, when compared with AGL1, EHA105 gained a larger number of hygromycin-resistant colonies per MYG plate for transformation (Figure 3A,B), indicating that A. tumefaciens strain EHA105 would help to improve the transformation efficiency
The results showed that themost that the 1/4 sawdust medium used for culturing dikaryotic strain W1 could produce the sawdust medium used for culturing dikaryotic strainpositive

Summary

Introduction

Filamentous fungi encompass diverse groups and are widely distributed in nature. Basidiomycetes, an indispensable member in filamentous fungi, play important roles in agriculture, industry, and medicine [1]. The genomes of several species have been sequenced and published, such as Coprinosis cinerea, Schizophyllum commune, Agaricus bisporus, Ganoderma lucidum, Lentinula edodes, etc. The establishment of genetic transformation method is crucial for the functional genomics research in filamentous fungi. The transformation method has been developed in several types of fungi, a highly efficient and convenient transformation system is desperately needed in Lentinula edodes. Present work established the Agrobacterium-mediated transformation (ATMT) of basidiomycete L. edodes in both monokaryon and dikaryon mycelia by using constructed binary plasmid pCAMBIA-1300-GFP. The transformation efficiency of ATMT was evaluated by using different mediums for recipient incubation and different varieties of L. edodes. While in the monokaryon strain W1-26, only the millet medium group obtained positive transformants with a positive frequency of 75.48%.

Results

Discussion

Conclusion