Mycopath (2014) 12(2): 77-82 Research Article Isolation of a putative invertase gene from the xerophilic Aspergillus niger GH1 strain F. Veana 1 , D. G. Rodríguez-Reyna 2 , E. T. Aréchiga-Carvajal 2 , C. N. Aguilar 1 , * R. Rodríguez-Herrera 1 1 Department of Food Science and Technology, School of Chemistry, Autonomous University of Coahuila. Saltillo, 25280, Coahuila, México. 2 Microbiology and Immunology Department. School of Biology. Autonomous University of Nuevo Leon. San Nicolás de los Garza, 66450, Nuevo León, México. * Corresponding author’s email: raul.rodriguez@uadec.edu.mx, rrh961@hotmail.com Abstract Invertase catalyzes the hydrolysis of sucrose into glucose and fructose and represents a biocatalyst of great importance in food industry. Some xerophilic fungal strains have a great potential to increase enzymatic production. In the present study, five xerophilic fungal strains (Penicillium pinophilum EH2, P. purpurogenum GH2, P. citrinum ESS, Aspergillus niger GH1 and A. fumigatus GS) previously reported as invertase producers were used for isolation and cloning of an invertase gene. A fragment of 1770 bp was isolated and cloned, which corresponding to invertase gene from A. niger GH1. On the other hand, multi- banding was observed from invertase gene from other strains and invertase gene could not be isolated. Results suggested that invertase gene and amino acid sequence exhibited 97% and 99% similarity to invertase from A. niger B60 and A. kawachii IFO 4308, respectively. Therefore, A. niger GH1 could be a good alternative for invertase production. Key words: β-D-fructofuranosidase, cloning, fructooligosacharides, Aspergillus niger, Penicillium. Introduction Invertases or β-fructofuranosidases (Ffase) (EC 3.2.1.26) catalyze the hydrolysis of terminal non-reducing β-D-fructofuranoside residues in β- D-fructofuranosides, including hydrolysis of sucrose into glucose and fructose. These enzymes are used in beverage and confectionary industries (Kumar et al., 2001). In addition, invertase also catalyze synthesis of fructo-oligosaccharides (FOS), which are promising ingredients for functional foods since they act as “prebiotics”, and exert a beneficial effect on human health, participating in the prevention of cardiovascular diseases, colon cancer, and osteoporosis (Linde et al., 2009; Kurakake et al., 2010). Industrially, invertases have been produced in submerged culture (SmC) using Saccharomyces cerevisiae. However, filamentous fungi, particularly Aspergillus niger strains secrete large amounts and varieties of enzymes, including invertase. In recent years, different xerophilic fungal strains were isolated from Mexican semi-desert and characterized as invertase producers (Veana et al., 2011; Flores-Gallegos et al., 2012). However, their nucleotide sequences have not been determined. Since, Boddy et al. (1993) described the sequence of invertase gene from A. niger B60, few studies have been reported about the use of heterologous expression system for increase the productivity of this enzyme. Present study was designed to analyze and describe the invertase nucleotide and amino acid sequence isolated from A. niger GH1. In addition, genetic relationship with other sequences of invertase was established. Materials and Methods Xerophilic fungal strains, host and vectors Penicillium purpurogenum GH2, P. pinophilum EH2, P. citrinum ESS, A. niger GH1 and A. fumigatus GS from the fungal collection of the Food Research Department-School of Chemistry-UAdeC (Cruz-Hernández et al., 2005) were used in this study. The pTOPO vector (10 ng μL -1 ) of TOPO TA cloning kit with pCR 2.1- TOPO, the pUC19 vector (10 pg μL -1 ) and electrocompetent Escherichia coli TOP10 cells were purchased from Invitrogen (San Diego, CA). PDA and LB agar Miller media were acquired from DIXON and US Biological, México. Tween 80 (Amresco, Ohio) and Wizard® SV Gel and PCR Clean-Up System from Promega (Madison, WI) were used. All chemicals were of analytical grade and purchased from Sigma-Aldrich Co. (St. Louis, MO) or from Productos Químicos Monterrey (Monterrey, Nuevo León, México).