title: New E. coli cloning vector using a cellulase gene (celA) as a screening marker.
abstract: The extracellular endoglucanase A gene of Clostridium thermocellum (celA) was used as a screening marker for E. coli cloning vector A 1.4-kb EcoRI fragment containing celA from pTvec/celA was isolated and cloned into a pUC18 deleting beta-galactosidase gene fragment. The constructed vectors, pCEL1, pCEL10, pCEL11, and pCEL20, have different multiple cloning sites within celA. If the cellulase, CelA, is inactivated by insertion of a foreign DNA fragment into multiple cloning sites, the recombinant transformants show no clear halos on an agar plate containing cellulose. This process overcomes the ambiguity of color screening in the X-gal/beta-galactosidase system, and over 90% of the recombinant transformants with no halos have foreign DNA inserts. Several E. coli strains were transformed successfully with pCEL series vectors regardless of mutation for alpha-complementation. Because E. coli strains do not have a cellulase gene, a vector using a cellulase gene screening marker can be used in any E. coli strain without limit. The new cloning system is very efficient, convenient, and cost effective.
authors.affiliation: Kyung Hee University, Suwon, Korea.
authors.forename: S J
authors.initials: SJ
authors.lastname: Jang
authors.forename: W J
authors.initials: WJ
authors.lastname: Park
authors.forename: S K
authors.initials: SK
authors.lastname: Chung
authors.forename: C Y
authors.initials: CY
authors.lastname: Jeong
authors.forename: D K
authors.initials: DK
authors.lastname: Chung
mesh_terms: D001483:Base Sequence; D002480:Cellulase / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics*; D005819:Genetic Markers; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D014169:Transformation, Bacterial
chemical_list: D005819:Genetic Markers; C074775:CelA endoglucanase; D002480:Cellulase