Characterization of a lytic Lactobacillus plantarum bacteriophage and molecular cloning of a lysin gene in Escherichia coli.
Bacteriophage SC921, which can infect Lactobacillus plantarum specifically, was isolated from a fermented vegetable source, Kimchi. This phage is active against six of 11 strains of L. plantarum tested as hosts. Morphologically, it has an isometric head (60 nm in diameter) and a non-contractile tail (260 nm long and 9-11 nm wide), indicating that it belongs to Bradley's group B or the Siphoviridae family according to the International Committee on Taxonomy of Viruses (ICTV). The bouyant density was 1.58 g/cm3. SDS-PAGE experimentation indicated that the phage particle contains two major structural proteins and several minor proteins. The genome was a double stranded linear DNA molecule with cohesive ends and 66.5 kb long by mapping genomic DNA digested with the restriction endonucleases: KpnI, SmaI, and XbaI. The [G + C] content of the phage DNA is 39.4%. For this lysin gene study, 9.4 kb of KpnI-digested DNA fragment was cloned into pUC19 and expressed in Escherichia coli. The KpnI fragment was considered as the genetic element responsible for the lysis gene of L. plantarum bacteriophage. The cloned fragment in pUC19 was hybridized to a 9.4-kb fragment generated by KpnI digestion of SC 921 as a probe. This confirmed that the fragment in pUC19 originated from phage DNA. The lysin gene was near the middle of the phage genome.
Yoon
S S
SS
Department of Biological Resources and Technology, College of Arts and Sciences, Yonsei University, Wonju, Kangwon-Do, South Korea. sungsik@dragon.yonsei.ac.kr
Kim
J W
JW
Breidt
F
F
Fleming
H P
HP
D001435:Bacteriophages / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000502:physiology*; D007778:Lactobacillus / Q000502:physiology; D009088:Mucoproteins / Q000235:genetics*
D004279:DNA, Viral; D009088:Mucoproteins; C010393:lysin, gastropoda