Thesis

Investigation of Various Transformation Methods to Knockout the gad Gene in Rhizopus oligosporus to Develop GABA-Free Tempe for Potential Comparative Zebrafish Behavior Study

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Tempe, a fermented soybean product from Indonesia, showcases how fermentation enhances
nutritional value and digestibility. Rhizopus oligosporus, a key starter culture in tempe production,
enzymatically enhances carbohydrate, vitamin, and mineral availability from raw soybeans. This
process also boosts bioactive compounds such as γ-aminobutyric acid (GABA), recognized for its role
as an inhibitory neurotransmitter that regulates movement and activity levels. GABA in tempe is
known to give a calming effect. Hence, the initial step to investigate it is to create a GABA-free strain,
involved knocking out the gad gene that is responsible for GABA production in R. oligosporus using
common fungal transformation methods, starting from the most convenient one to the traditional
one, which are PEG-mediated, electroporation, and agrobacterium-mediated approaches.
Unfortunately, none of the methods tested successfully knocked out the gad gene, likely due to the
fungus's thick cell wall hindering nucleic acid insertion. Future research should explore novel
protocols and enzymes to generate fungal protoplasts for further investigations."
"Tempe, a fermented soybean product from Indonesia, showcases how fermentation enhances
nutritional value and digestibility. Rhizopus oligosporus, a key starter culture in tempe production,
enzymatically enhances carbohydrate, vitamin, and mineral availability from raw soybeans. This
process also boosts bioactive compounds such as γ-aminobutyric acid (GABA), recognized for its role
as an inhibitory neurotransmitter that regulates movement and activity levels. GABA in tempe is
known to give a calming effect. Hence, the initial step to investigate it is to create a GABA-free strain,
involved knocking out the gad gene that is responsible for GABA production in R. oligosporus using
common fungal transformation methods, starting from the most convenient one to the traditional
one, which are PEG-mediated, electroporation, and agrobacterium-mediated approaches.
Unfortunately, none of the methods tested successfully knocked out the gad gene, likely due to the
fungus's thick cell wall hindering nucleic acid insertion. Future research should explore novel
protocols and enzymes to generate fungal protoplasts for further investigations.

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Availability

Detail Information

Series Title

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Call Number

BT 24-002

Publisher

i3L, Jakarta : i3L Press., 2024

Collation

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Language

English

ISBN/ISSN

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Classification

NONE

Content Type

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Media Type

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Carrier Type

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Edition

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Subject(s)

tempe
Fungal transformation
Rhizopus oligosporus
γ-aminobutyric acid (GABA)

Specific Detail Info

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Statement of Responsibility

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