Genome editing vector, genome editing system constituted by genome editing vector and application

Abstract

The invention relates to a genome editing vector, a genome editing system constituted by the genome editing vector and an application, and in particular relates to a thermophilic fungi genome DNA editing vector, a genome editing system CRISPR / Cas9 constituted by the genome editing vector as well as an editing method an application. The genome editing vector comprises a promoter for promoting coding DNA transcription of sgRNA, wherein the promoter is an RNA polymerase III type U6 promoter. With the application of the genome editing system, genome editing efficiency of myceliophthora strains M.thermophile and M.heterothallica can be obviously improved, and simultaneous editing of multiple sites of myceliophthora genome can be implemented, so that polygenic mutant strains are obtained; the series of mutant strains can obviously improve production capacity of cellulase and can be applied to transformation of thermophilic fungi genetic engineering for high yield of the cellulase; the genomeediting system can also promote researches on gene functions of myceliophthora thermophila; and meanwhile, the genome editing system is of significance to genome directional editing and metabolic engineering transformation of thermophilic industrial cellulase production strains.

Description

technical field [0001] The invention belongs to the field of genetic engineering and biotechnology, and relates to the development of a CRISPR / Cas9 system, in particular to a genome editing carrier, its composed genome editing system and its application, especially to a thermophilic fungus genome DNA editing carrier, its The composed genome editing system CRISPR / Cas9 and editing methods and applications. Background technique [0002] Myceliophthora is a thermophilic filamentous fungus that can rapidly degrade cellulose. It has a fast metabolic rate and can secrete a large number of lignocellulolytic enzymes. The types and quantities of its enzyme systems are quite rich. Compared with the industrial cellulase producing bacteria Trichoderma reesei and Penicillium decumbens, the high-temperature fermentation of Myceliophthora thermophila and the enzyme system produced have the advantages of high activity and high stability at high temperature, and become natural high-temperatur...

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Problems solved by technology

[0005] Although the CRISPR / Cas9 system has been successfully developed in some mesophilic fungi, whether the CRISPR / Cas9 system can work in thermostable filamentous fungal cells, especially whether it is possible to find highly efficient inducible guide RNA (gRNA) in thermophilic fungi Whether the RNA polymerase type III promoter expressed in vivo, including Cas9 and gRNA, can be introduced into Myceliophthora thermophila cells through in vivo transcription to successfully edit the target site, and whether it can specifically target the genomic DNA of the thermophilic fungus , so far these studies have not been reported and explained

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