Detection of pathogenic bacteria by using zinc finger protein fused with firefly luciferase.
We constructed a novel bacterial genome detection system using zinc finger protein (ZF) fused with firefly luciferase (ZF-luciferase). Taking advantage of the direct recognition of double-stranded DNA (dsDNA) by ZF, we previously constructed bacterial genome detection systems that did not require dehybridization processes. To detect polymerase chain reaction (PCR) products rapidly and with a high sensitivity, we constructed two kinds of ZF-luciferase, Sp1-fused luciferase (Sp1-luciferase), and Zif268-fused luciferase (Zif268-luciferase). ZF-luciferase not only maintains luciferase activity but also shows dsDNA-binding ability and specificity. Furthermore, we succeeded in the detection of 10 copies of the genome of Legionella pneumophila and Escherichia coli O157. ZF-luciferase would be a useful tool for highly sensitive detection of pathogenic bacterial genome.
Department of Biotechnology and Life Science, Tokyo University of Agriculture & Technology, Koganei, Japan.
Koichi
K
Abe
Takenori
T
Kumagai
Chiaki
C
Takahashi
Aki
A
Kezuka
Yoshiyuki
Y
Murakami
Yuko
Y
Osawa
Hiroaki
H
Motoki
Takafumi
T
Matsuo
Michio
M
Horiuchi
Koji
K
Sode
Shizunobu
S
Igimi
Kazunori
K
Ikebukuro
D001665:Binding Sites; D004269:DNA, Bacterial / Q000032:analysis; D051766:Early Growth Response Protein 1 / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000302:isolation & purification*; D016952:Legionella pneumophila / Q000235:genetics / Q000302:isolation & purification*; D049409:Luciferases, Firefly / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D016335:Zinc Fingers / Q000235:genetics*
D004269:DNA, Bacterial; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D011993:Recombinant Fusion Proteins; D016329:Sp1 Transcription Factor; D049409:Luciferases, Firefly