Cloning and amplification of the .beta.-glucosidase gene of Trichoderma reesei
AssigneeGenencor International, Inc.
InventorsTimothy Fowler, Christopher C. Barnett, Sharon Shoemaker
Life of this patent
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Abstract
A process for expressing extracellular .beta.-glucosidase in a filamentous fungus by expressing a fungal DNA sequence encoding enhanced, deleted or altered .beta.-glucosidase in a recombinant host microorganism is disclosed. Recombinant fungal cellulase compositions containing enhanced, deleted or altered expression of .beta.-glucosidase is also disclosed.
Claims19
A process for expressing extracellular .beta.-glucosidase from DNA encoding an extracellular .beta.-glucoidase which DNA is obtained from a microorganism selected from the group consisting of Trichoderma, Aspergillus, and Neurospora, and wherein said DNA sequence or a portion thereof is capable of amplification by PCR with SEQ ID NO:
- 3 and SEQ ID NO:
- 4, wherein the amplification conditions are denaturation at 95.degree. C. for 10 minutes, annealing at 50.degree. C. for 2 minutes and extension at 65.degree. C. for 10 minutes for 30 cycles and wherein the amplification product is a DNA sequence that comprises a DNA sequence of about 700 base pairs from about position 471 to about position 1171 of SEQ ID NO:
- 1, comprising:
- expressing said extracellular .beta.-glucosidase from a DNA insert through recombinant techniques in a filamentous fungus, wherein the inserted DNA comprises all of the coding region of a fungal .beta.-glucosidase gene and sequence necessary for the .beta.-glucosidase gene's transcription and translation, and wherein said filamentous fungus is selected from the group consisting of Trichoderma, Aspergillus, Neurospora, Humicola and Penicillium.
A process for expressing extracellular .beta.-glucosidase, comprising a) inserting DNA encoding an extracellular .beta.-glucosidase gene obtained from a microorganism selected from the group consisting of Trichoderma, Aspergillus, and Neurospora, wherein said DNA sequence or a portion thereof is capable of amplification by PCR with SEQ ID NO:
- 3 and SEQ ID NO:
- 4, wherein the amplification conditions are denaturation at 95.degree. C. for 10 minutes, annealing at 50.degree. C. for 2 minutes and extension at 65.degree. C. for 10 minutes for 30 cycles and wherein the amplification product is a DNA sequence that comprises a DNA sequence of about 700 base pairs from about position 471 to about position 1171 of SEQ ID NO:
- 1, and wherein said host fungus is selected from the group consisting of Trichoderma, Aspergillus, Neurospora, Humicola and Penicillium;
- b) isolating transformants having enhanced .beta.-glucosidase expression as compared to a non-transformed filamentous host;
- c) culturing said transformants under conditions to permit growth of said transformants;
- and d) isolating a fungal cellulase composition containing the .beta.-glucosidase from said transformants.
A probe for use in detecting nucleic acid sequences encoding .beta.-glucosidase from a filamentous fungi comprising SEQ ID. NO:
- 1.
A polynucleotide sequence comprising the nucleic acid sequence of SEQ ID NO:
- 1.
Dependencies are inferred from the claim text.