Transformation of Saccharopolyspora erythraea by electroporation of germinating spores: construction of propionyl Co-A carboxylase mutants

Autor:	J S Lampel, R S English, TJ Vanden Boom
Rok vydání:	1998
Předmět:	biology Electroporation Mutant Germ tube Propionyl-CoA carboxylase Bioengineering biology.organism_classification Applied Microbiology and Biotechnology Transformation (genetics) Biochemistry Spore germination Saccharopolyspora erythraea Biotechnology Transformation efficiency
Zdroj:	Journal of Industrial Microbiology and Biotechnology. 21:219-224
ISSN:	1476-5535 1367-5435
DOI:	10.1038/sj.jim.2900576
Popis:	The introduction of plasmid DNA into germinating spores of an industrially improved strain of Saccharopolyspora erythraea was accomplished by electroporation. Various parameters affecting the efficiency of electroporation were examined. The most critical factor was the extent of spore germination. Electrocompetence was limited to a 4-h period following the initial emergence of the germ tube. Electroporation efficiencies as high as 2 × 105 CFU μg−1 of plasmid DNA were obtained using electrocompetent germlings. The optimal field strength was 12–14 kV cm−1 with a pulse duration of 15–20 ms. Electrocompetent germlings were stored at −80°C without a significant decrease in transformation efficiency. The utility of this protocol was demonstrated by isolating a propionyl-CoA carboxylase mutant through targeted gene disruption and replacement.
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=doi_________::a26d862d18b65d39551a5be935d852cc https://doi.org/10.1038/sj.jim.2900576 Zobrazit plný text záznamu Plný text ve formátu PDF