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Updated: Jul 6, 2026

Transfection, Selection, and Colony-picking of Human Induced Pluripotent Stem Cells TALEN-targeted with a GFP Gene into the AAVS1 Safe Harbor

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A highly efficient gene-targeting system for Aspergillus parasiticus.

¹Southern Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, New Orleans, LA 70124, USA. pkchang@srrc.ars.usda.gov

Letters in Applied Microbiology

|March 19, 2008

Summary

A new gene-targeting system in Aspergillus parasiticus significantly boosts efficiency by disrupting the NHEJ pathway. This method achieves 96% targeting frequency, accelerating functional genomic studies.

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Genetic Manipulation in Δku80 Strains for Functional Genomic Analysis of Toxoplasma gondii

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Published on: July 12, 2013

Area of Science:

Microbiology
Molecular Biology
Genetics

Background:

Aspergillus parasiticus is a significant producer of aflatoxins.
Efficient gene targeting is crucial for understanding fungal genomics and developing control strategies.

Purpose of the Study:

To develop a highly efficient gene-targeting system for Aspergillus parasiticus.
To increase the frequency of targeted gene replacement and disruption.

Main Methods:

Disruption of the ku70 gene, involved in the nonhomologous end-joining (NHEJ) pathway, to create an NHEJ-deficient strain (RHDeltaku70).
Manipulation of flanking regions of the aflatoxin biosynthetic gene (adhA) to create homologous ends for integration.
Utilization of a linearized DNA fragment with a pyrithiamine resistance (ptr) marker for selection.

Main Results:

The NHEJ-deficient strain (RHDeltaku70) exhibited normal growth and development.
Targeting the adhA gene in RHDeltaku70 achieved a remarkable 96% gene-targeting frequency.
The homologous recombination pathway was confirmed as the primary DNA repair mechanism in A. parasiticus.

Conclusions:

The developed system, combining NHEJ deficiency, simplified homologous end creation, and ptr-based selection, offers a highly efficient gene-targeting method.
This system significantly reduces the time and effort required for generating knockout strains.
The system is applicable for targeted gene replacement, disruption, and modification of specific chromosomal locations, accelerating functional genomic studies in Aspergillus species.