Influence of Bacteriophage Lambda Gene P Expression On Host Escherichia coli Cells
Date
2011-11-18
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
ORCID
Type
Degree Level
Masters
Abstract
Bacterial viruses have been an important tool for molecular biological discoveries.
Bacteriophage is a bacterial virus that has been intriguing researchers for over five decades.
But still, we are yet to answer many questions about bacteriophage . genes, O and P play an
important role in replication initiation. P outcompetes host DnaC and recruits and directs
DnaB helicase to the origin of replication, ori .
My study shows that P expression is lethal to the host cell. The P-survivor cells were
examined and were found to have chromosomal mutations. This led to the possibility that P
protein may be involved in elevating the level of random mutations of the host chromosome.
These studies explore this possibility. The influence of P expression on the appearance of
chromosomal mutations was assessed by using rifampicin resistance, and auxotrophy as
chromosomal targets. Cellular expression of P from a cryptic prophage or from a ColE1-type
plasmid was employed in this study.
Insertional inactivation of P by recombineering knocked out P-lethality and the potential
mutator phenotype among the 42°C survivors. The apparent mutator phenotype was also lost in
42°C survivors upon induction of a cryptic prophage with wild-type O and an insertion in P.
When wild-type gene P on a ColE1 plasmid was replaced by a deleted P or a mutated P (P )
gene, the rate of rifampicin resistance among the 37°C cfu was significantly lowered. These
observations suggest that there might be a relation between P expression and the appearance of
mutations among the P-survivors. My data also support and extend laboratory findings that two
missense mutations in host dnaB knock out P-lethality and suppress the observed potential
mutator phenotype.
ColE1 plasmid loss occurred among the P-survivor cells when P was expressed from a
ColE1 plasmid in a wild-type cell. My data suggested that the plasmid-less or cured rifampicin
resistant mutants that arose upon thermal induction of P and the 594 rifampicin resistant mutants
that arose in the absence of P expression were resistant to P-toxicity. However, my data showed
that a 9 base pair deletion in the rpoB gene (mutation in rpoB confers rifampicin resistance
iii
phenotype to the cells) that affected 4 amino acids was sensitive to P-toxicity which proved that
the deletion itself did not make the cells resistant to P-toxicity.
Description
Keywords
Bacteriophage Lambda, Escherichia coli, ColE1 plasmid, potential mutator phenotype, rifampicin resistance, auxotrophy, P-lethality.
Citation
Degree
Master of Science (M.Sc.)
Department
Microbiology and Immunology
Program
Microbiology and Immunology