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Identification and Characterization of a Neutral Locus for Knock-in Purposes in C. parapsilosis

Authors: Tibor Nemeth; Csaba Papp; Csaba Vagvolgyi; Tanmoy Chakraborty; Attila Gacser; Attila Gacser;

Identification and Characterization of a Neutral Locus for Knock-in Purposes in C. parapsilosis

Abstract

Invasive fungal infections caused by Candida species affect approximately 700,000 people worldwide resulting in 300,000 deaths annually. Besides Candida albicans, other members of the genus have gained relevance in the last two decades, including C. parapsilosis whose incidence is particularly high amongst low birth weight neonates. To investigate the virulence properties of this pathogen several techniques have been developed for generating knock-out mutants, however, no target locus for knock-in approaches have been published so far. Here we report CpNEUT5L (N5L), an intergenic locus in C. parapsilosis, and introduce an integrative GatewayTM and a classical ligation based replacement plasmid to target it with. As a proof of principle, we fluorescently tagged laboratory and prototroph strains and established that this locus is also suitable for reintegration purposes. We concluded that GFP-expressing constructs integrated into this region provide strong, homogenous fluorescent signals while alteration of this locus affects neither the growth of the mutants in liquid or on solid media, even in the presence of different stressors, nor their basic virulence properties. Hence, our findings demonstrate that N5L is a highly effective neutral locus for knock-in approaches in C. parapsilosis.

Country
Hungary
Keywords

Candida parapsilosis, reintegration, GFP, knock-in, Microbiology, QR355 Virology / víruskutatás, genetic manipulation, QR1-502, overexpression

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    popularity
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    Top 10%
    influence
    This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
    Average
    impulse
    This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
10
Top 10%
Average
Top 10%
Green
gold