GENETIC ENGINEERING OF FUNGAL STRAINS USING CRISPR/Cas9 MULTIPLEX TO PRODUCE TAILOR-MADE CELLULOLYTIC COCKTAILS

• Grant number: 17/26370-0
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• Start date: April 03, 2018
• End date: April 02, 2019
• Field of knowledge: Biological Sciences - Microbiology - Applied Microbiology
• Principal Investigator: André Ricardo de Lima Damasio
• Grantee: Fabiano Jares Contesini
• Supervisor: Uffe Hasbro Mortensen
• Host Institution: Instituto de Biologia (IB). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil
• Institution abroad: Technical University of Denmark (DTU), Denmark
• Associated to the scholarship: 17/10083-1 - GENETIC ENGINEERING OF A FUNGAL PLATFORM TO SECRETE LIGNOCELLULOLYTIC ENZYMES FOR OLIGOSACCHARIDE PRODUCTION, BP.PD

Abstract

Complete degradation of plant biomass using enzymes is a key step for biofuel production inthe biorefinery context. The assimilation of oligosaccharides by fermentative microorganisms,instead of monosaccharides, is considered advantageous since it can decrease the competitionwith possible contaminants, resulting in improvements regarding process yield andproductivity. For this purpose, apart from the necessity of engineered yeasts capable ofinternalizing xylo- and cello-oligosaccharides, an efficient tailor-made cellulolytic cocktail isrequired. Filamentous fungi, such as Aspergillus spp., are excellent candidates for thedevelopment of platforms for enzyme production, mainly due to their large capability ofprotein secretion. In this context, the main objective of this proposal is to implementCRISPR/Cas9 multiplex systems in Aspergillus niger and Aspergillus nidulans aiming toinactivate genes coding for cellobiohydrolases, ²-glucosidases and ²-xylosidases. Theseenzymes were selected since they are responsible for the formation of monosaccharides.CRISPR/Cas9 multiplex is an innovative system that allows inactivation of a series of specificgenes by truncation or deletion. After implementing the technique in both Aspergillus species,we will perform one or more rounds of gene inactivation in order to obtain improved fungalstrains producing tailor-made enzymatic cocktails that can partially degrade plant biomassenriching the products in terms of oligosaccharides, compared to the parental strains. Weexpect to learn this technology with the supervision of Prof. Uffe H. Mortensen in theDepartment of Biotechnology and Biomedicine at the Technical University of Denmark (DTU),who is an expert in CRISPR/Cas9 technology in Aspergillus, a technology that has not beendeveloped in Brazil yet.