Molecular tools for gene manipulation in filamentous fungi

Abstract

Functional genomics of filamentous fungi has gradually uncovered gene information for constructing 'cell factories' and controlling pathogens. Available gene manipulation methods of filamentous fungi include random integration methods, gene targeting technology, gene editing with artificial nucleases and RNA technology. This review describes random gene integration constructed by restriction enzyme-mediated integration (REMI); Agrobacterium-mediated transformation (AMT); transposon-arrayed gene knockout (TAGKO); gene targeting technology, mainly about homologous recombination; and modern gene editing strategies containing transcription activator-like effector nucleases (TALENs) and a clustered regularly interspaced short palindromic repeat/associated protein system (CRISPR/Cas) developed in filamentous fungi and RNA technology including RNA interference (RNAi) and ribozymes. This review describes historical and modern gene manipulation methods in filamentous fungi and presents the molecular tools available to researchers investigating filamentous fungi. The biggest difference of this review from the previous ones is the addition of successful application and details of the promising gene editing tool CRISPR/Cas9 system in filamentous fungi.

Cys 2 His 2 Zinc Finger Transcription Factor BcabaR1 Positively Regulates Abscisic Acid Production in Botrytis cinerea

“…Functional analysis of bcabaR1 was performed by gene silencing and overexpression, followed by gene expression analysis in strain TB-31, and the results revealed that bcabaR1 affected ABA production by regulating the ABA-biosynthetic gene cluster. In the filamentous fungus, RNA-mediated gene silencing is a reliable genetic manipulation strategy in analyzing gene function (29). Patel et al (30) first reported RNA silencing of a superoxide dismutase gene in B. cinerea, and the functions of several other B. cinerea genes that participate in pathogenicity or other biological processes have since been investigated using RNA interference (31)(32)(33).…”

Advances in targeting and heterologous expression of genes involved in the synthesis of fungal secondary metabolites

“…29 A disadvantage of ATMT is that it is time-consuming because it requires optimization of transformation conditions, such as optimization of ratio of host cells to the bacteria, co-cultivation temperature, and co-cultivation time. 12 ATMT has successfully targeted genes in a variety of fungi. Ustilaginoidea virens causes rice false smut and harms the nervous system of animals.…”

“…Previous reviews have summarized many techniques for fungal genetic studies, but they did not pay much attention to single gene or single BGC manipulation techniques. 11,12 This review focused on recent progress in methods specically for individual fungal gene or fungal BGC targeting and heterologous expression, as exemplied by ergot alkaloid biosynthesis.…”

Rhodosporidium toruloides - A potential red yeast chassis for lipids and beyond

“…Moreover, as the intrinsic feature of ATMT transformation is random integration of the genome, it is essential to screen massive clones to identify those with superior phenotype and to diminish the risk of unexpected interference with endogenous genes (Wang et al . 2017 ).…”