Article

Transgenic Research

, Volume 2, Issue 2, pp 84-92

First online:

Glucoamylase overexpression inAspergillus niger: Molecular genetic analysis of strains containing multiple copies of theglaA gene

  • Jan C. VerdoesAffiliated withDepartment of Molecular Genetics and Gene-Technology, TNO Medical Biological Laboratory
  • , Peter J. PuntAffiliated withDepartment of Molecular Genetics and Gene-Technology, TNO Medical Biological Laboratory
  • , Jaap M. SchrickxAffiliated withDepartment of Microbiology, Vrije Universiteit, Biological Laboratory
  • , Henk W. van VerseveldAffiliated withDepartment of Microbiology, Vrije Universiteit, Biological Laboratory
  • , Adriaan H. StouthamerAffiliated withDepartment of Microbiology, Vrije Universiteit, Biological Laboratory
  • , Cees A. M. J. J. van den HondelAffiliated withDepartment of Molecular Genetics and Gene-Technology, TNO Medical Biological Laboratory

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Abstract

A strategy, based on the usage of theamdS selection marker and a cosmid vector containing four copies of the glucoamylase gene (glaA), was developed to obtain glucoamylase (GLA)-overproducingA. niger strains. With this strategy, fungal strains carrying up to 200 copies of theglaA gene could be isolated at a relatively high frequency. In each transformant analysed, integration occurred in a single chromosome. A significant increase in the extracellular GLA production was observed in most of the transformants carrying multiple copies of theglaA gene. Further analysis showed that the amount of GLA that is produced was not proportional to the number ofglaA copies in these transformants. However, the level of GLA production clearly correlated with the amount ofglaA mRNA produced in these transformants. From these results it is concluded that GLA production is limited at the level of transcription.

Keywords

transcription regulation regulatory protein amdS selection marker cosmid vector CHEF analysis