Generating genomic platforms to study Candida albicans pathogenesis

Mélanie Legrand; Sophie Bachellier-Bassi; Keunsook K. Lee; Yogesh Chaudhari; Hélène Tournu; Laurence Arbogast; Hélène Boyer; Murielle Chauvel; Vitor Cabral; Corinne Maufrais; Audrey Nesseir; Irena Maslanka; Emmanuelle Permal; Tristan Rossignol; Louise A. Walker; Ute Zeidler; Sadri Znaidi; Floris Schoeters; Charlotte Majgier; Renaud A. Julien; Laurence Ma; Magali Tichit; Christiane Bouchier; Patrick Van Dijck; Carol A. Munro; Christophe d’Enfert

Construction of C. albicans UME6-overexpression strains. Detailed methods for the transfer of C. albicans ORFs from pDONR207 into the expression plasmids as well as the integration of the resulting expression plasmids at the RPS1 locus have been described (34) . Briefly, the UME6 ORF was transferred from the entry clone into each one of 15 expression vectors by using the Gatewayâ„¢ LR Clonaseâ„¢ II Enzyme Mix (Invitrogen). After E. coli transformation, the plasmids were verified by EcoRV digest. The URA3-and NAT1-bearing expression plasmids were digested by StuI and transformed into C. albicans strain CEC161 or CEC369, respectively, according to Walther and Wendland (39) . Transformants were selected for prototrophy or nourseothricin resistance, respectively, and verified by PCR using primer CIpUL with (i) primer CIpUR for the URA3-bearing plasmids and (ii) primer author/funder. All rights reserved. No reuse allowed without permission.