Construction of MYC-tagged Sch9p in Candida albicans by using of long primers PCR amplification

Abstract

Abstract:

Objective To construct the MYC-tagged Sch9p in Candida albicans.Methods Using a pair of the long primers to amplify sequences containing the MYC tag and the ARG4 selection markers from the plasmid pFA-ARG4-MYC.The amplified plasmid sequences were transformed into the C-terminus of SCH9 open reading frame (ORF) in C.albicans SN152 by homologous recombination.The positive colonies were selected in the SC-Leu^- selective solid culture medium.Afterwards,the positive colonies with correct integration were confirmed by PCR of genomic DNA.Finally,these positive transfectants were examined by time-growth curve testing,drug sensitivity spot assays,and mycelium inducing experiments.Results Strains with MYC-tagged Sch9p which had normal phenotypes were selected.Conclusions The MYC-tagged Sch9p of C.albicans strains can be constructed correctly by using of long primers to amplify the plasmid sequences containing the MYC tag and ARG4 selection markers which were integrated into the C-terminus of SCH9 ORF in C.albicans by homologous recombination.

Key words: Candida albicans, MYC, SCH9, long primer

CLC Number:

R379.4

XU Qiu-rong, LV Quan-zhen, SUI Xue, WANG Xiao-juan, YAN Lan, JIANG Yuan-ying. Construction of MYC-tagged Sch9p in Candida albicans by using of long primers PCR amplification[J]. Chinese Journal of Mycology, 2015, 10(3): 129-133.

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