Abstract

A strain improvement program was initiated to increase the yield of extracellular phytase (EC 3.1.3.8) in Aspergillus ficuum NRRL 3135. A minimal plating medium containing 2.99 g liter-1 a-ketoglutaric acid was developed to decrease and stabilized conidiogenesis time and facilitate the isolation of single colonies. Exposure of A. ficuum conidia to 0.5 mg ml-1 N-methyl-N’-nitro-N-nitrosoguanidine (NTG) for 15 min at pH 6.5 caused a one log kill and an increase in the mutation rate of 800-fold. Treatment of A. ficuum conidia with 46 ergs mm-2 sec -1 UV radiation for 30 sec resulted in a one log kill and a 100,000-fold increase in mutation rate. Mutants which overproduce extracellular acid phosphatase(s) were tentatively identified in a plate assay by histochemical staining with 0.1 g ml-1 a-naphthyl acid phosphate and 0.04 g ml-1 fast black K-salt in 0.2 M acetate buffer pH 4.8. Confirmation of extracellular acid phosphatase overproduction was by analysis of shaker flask fermentation broths via colorimetric enzyme assays with 0.04 M p-nitrophenyl phosphate and 2.5 µM sodium phytate as substrates. A UV derived mutant had a two-fold decrease in conidiogenesis time and a 17% increase in acid phosphatase activity. Further characterization of this mutant by SDS-PAGE electrophoresis and inhibition studies using 1 mM fluoride showed that the apparent increase in enzyme activity was caused by hyperproduction or hypersecretion of a nonspecific acid phosphate (EC 3.1.3.2).

Notes

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Graduation Date

1987

Semester

Spring

Advisor

Wodzinski, Rudy J.

Degree

Master of Science (M.S.)

College

College of Arts and Sciences

Degree Program

Microbiology

Format

PDF

Pages

55 p.

Language

English

Rights

Public Domain

Length of Campus-only Access

None

Access Status

Masters Thesis (Open Access)

Identifier

DP0020546

Contributor (Linked data)

Rudy J. Wodzinski (Q60389297)

Accessibility Status

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