Journal
APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
Volume 170, Issue 6, Pages 1533-1545Publisher
SPRINGER
DOI: 10.1007/s12010-013-0297-9
Keywords
Penicillium citrinum; Protoplast fusion; Nuclease P1; Sporulation; Genome shuffling
Funding
- National Nature and Science Foundation of China [3117175]
- Nature and Science Foundation of Zhejiang Province [Y3100609]
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Genome shuffling is a powerful approach for efficiently engineering industrial microbial strains with interested phenotypes. Here we reported a high producer of nuclease P1, Penicillium citrinum G-16, that was bred by the classical physics-mutagenesis and genome shuffling process. The starting populations were generated by Co-60 gamma-irradiation mutagenesis. The derived two protoplast fractions were inactivated by heat-treatment and ultraviolet radiation respectively, then mixed together and subjected to recursive protoplast fusion. Three recombinants, E-16, F-71, and G-16, were roughly obtained from six cycles of genome shuffling. The activity of nuclease P1 by recombinant G-16 was improved up to 1,980.22 U4/ml in a 5-l fermentor, which was 4.7-fold higher than that of the starting strain. The sporulation of recombinant G-16 was distinguished from the starting strain. Random amplified polymorphic DNA assay revealed genotypic differences between the shuffled strains and the wild type strain. The close similarity among the high producers suggested that the genetic basis of high-yield strains was achieved by genome shuffling.
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