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Abstract
Strain improvement is an important tool in commercial development of microbial fermentation processes for hyper production of enzymes. Aspergillus niger was isolated and identified as best pectinase producer through submerged fermentation and the same culture was subjected to mutate through physical (UV irradiation at 254 nm) and chemical (ethidium bromide, ethyl methyl sulphonate and sodium azide) mutation for enhancement of pectic acid-degrading enzymes (pectinase). Mutated A.nigerUV radiation exposure for 60 mins yields 260 nkat of pectinase activity when compared with wild strain yielded 235.06nkat. A mutant of A.niger showed higher enzyme activity on treatment with ethyl methyl sulphonate (2mg/ml for 60 mins) followed ethidium bromide (6mg/ml for 60mins) treatment found to be 265 nkat and 340 nkat of pectinase activity respectively. The pectinase producer which is susceptive to sodium azide (0.1% for 30min) yields 180 nKat pectinase activity. The combined UV and EtBr treatment (60min, 6mg/ml) yielded mutant with 1.69 fold enhanced polygalacturonase production compared with wild strain. The structure of poly galactouronase gene (pgal) was investigated and the amino acid sequence encoding enzyme was found to have 99% similarity with other A.niger isolates.
Key words: Aspergillus niger, mutation, ethidium bromide, sodium azide, pgal gene sequencing.
Introduction
For commercial production of enzyme, filamentous fungi are more commonly employed than yeast and bacteria [1]. Saprophytic and plant pathogenic fungi produces various enzymes for degrading plant cell wall component, which majorly comprises pectinase, the enzyme involved in the degradation of pectin. Pectinase are the group of enzymes involved in Pectin degradation; a polysaccharide substrate found in the cell wall and middle lamella of plants [2]. Pectins are high molecular weight, negatively charged, acidic, complex glycosidic macromolecules that are present in the Plant kingdom. Most pectic enzyme are used in the fruit processing industry. Pectic enzymes alone account for about one quarter of the world's food enzyme production [3]. The most common application of pectinase is in the food industry to extract and clarify fruit and beverages [4,5]. Pectinases are also used in industrial processes, such as in ramie fiber degumming, oil extraction, coffee and tea fermentation, and industrial wastewater treatment [6,7].
Production of Pectinase has been largely focused in Aspergillus niger[8]. A.niger is a work horse of industrial microbiology that has been subjected to different...