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GENE TRANSFER IN BACTERIA (Conjugation (Hfr Strains (Time dependent…
GENE TRANSFER IN BACTERIA
Reproduction in Prokaryotes
mostly
haploid
reproduce by
binary fission
:one: parent cell enlarges cell wall/ membrane and begins to replicate
:two: septum forms as chromosomes move to
cell end
:three:
Full septum divides cells
:arrow_right: separation to two daughter cells
:warning: genetic differences can only occur by
Mutation
bc there's no crossover occur (due to the asexual reproduction)
Horizontal
Gene Transfer
:pen: transmission of genetic material btwn different cells :arrow_right: exchanging genes within different species or same spcies
Transformation :arrow_right: cells take up
naked DNA
from their
environment
Conjugation :arrow_right:
cell to cell
transfer of DNA
Transduction :arrow_right:transfer mediated by
viruses
:warning: different from
sexual reproduction
bc
it doen't have chromosome paired up
Conjugation
Lederberg & Tatum
Used
auxotrophic
mutants for each specific strain
Strain A-
:arrow_forward: met-bio-thr+leu+thi+ :arrow_right: needs methionine and biotin to grow
Strain B-
:arrow_forward: met+bio+thr-leu-thi- :arrow_right: needs threonine, leucine and thiamine to grow
Plated these bacterial strains on agar
without these biosynthates
:cry:
individually
:arrow_right: No bacterial growth (no colonies)
:smiley:
together
:arrow_right: Some bacterial growth with
wildtype strain
(all plus)
:red_flag: suggesting that
there were some gene exchange here
:arrow_right: some form of
DNA Recombination
Hayes
:skull_and_crossbones:Treated these two strains with
streptomycin
(slowly
kill cell
)
On agar plate (
same as above
)
:check: killed
strain A
with streptomycin,
strain B
is alive :arrow_right:
Bacterial growth
:smiley:
:check: killed
strain B
with streptomycin,
strain A
is alive :arrow_right:
No bacterial growth
:cry:
:red_flag: suggesting that
genetic transfer was unidirectional
:arrow_right:
A :arrow_forward: B :arrow_forward: progeny
:point_right:Stain A is
donor
:arrow_right: donates DNA to Strain B (
recipient
)
Discovery of the Fertility Factor
(F)
Gene transfer is
unidirectional
:arrow_right: one is donor, the other one can only be recipients
:point_right: F+ is donor, F- is recipient
F is a
plasmid
:arrow_right: directs the
synthesis of pili
Plasmid
:arrow_right: small
circular
non-chromosomal
DNA element
Pilus
:arrow_right:
hair-like
appendage on bacterial surface :arrow_right:
connects two bacteria
:warning:
Plasmid can be integrated into chromosomes
:red_cross: If plasmid
NOT integrated
:arrow_right: only few colonies survive
:check:
an example of pre existing mutation in the population
:arrow_right: Mutation in this case is integration of
F into genome
:one: pilus from F+ pulls bacterial together
:two: An enzyme makes
single strand breakage
at
origin of transfer
(oriT) in
plasmid
:three:
plasmid encoded helicase
unwinds plasmid DNA :arrow_right:
broken strand is transferred to recipient
:four: DNA synthesis converts
s.s. DNA :arrow_forward: d.s. DNA
in
donor and recipient
:arrow_right: each cell have a copy of plasmid
Hfr Strains
:pen:
High frequency of recombination
strains :arrow_right: has F factor integrated into bacterial chromosome :arrow_right:
F+ with the integration of F factor into its chromosome
Exconjugant
of F- :arrow_right: cells have transferred fragment converting into double helix :arrow_right:
results in Recombinant
:star:
in F- :arrow_right: double crossover inserts a piece of plasmid to recipient chromosome
:no_entry:
very efficient to deliver genetic material (in linear chromosome) to F-(recipient)
:arrow_right:
F+ becoming Hfr :arrow_forward: F-
Time dependent transmission of Hfr genes
Each donor allele
first appears in the F- recipients
at
a
specific time
and
specific sequence
after mating began
Later donor alleles are present in fewer recipient cells
:+1:
the transfer process will stop before the furthermost genes are transferred
:arrow_right: they only included in few ricipients
F- rarely converted in Hfr or F+
:arrow_right: bc the part of F conferring
donor ability
transmitted with
low frequency
:red_flag:suggesting that
F factor is the last part of linear chromosome
:warning:Linked map :
O-a-b-c-F
Chromosome circularity
:arrow_right: F factor integration determines order of gene transfer in Hfr strains
Transformation
naked DNA
released from
dead
or
live
cells
Occur
naturally
or
artificially
Competent cell
able to
take up
DNA and be
transformed
:warning:
Most competent
during
exponential growth
:arrow_right: when it's dramatically increasing the # of
viable cells
artificially induced by :one: Chemical (CaCl) :two:Physical (electric shock)
:one: Competent cell binds naked DNA by a
protein complex
:two: one strand of naked DNA is
degraded
by
nuclease
:three: other strand is
transported
into competent cell
:four: The naked DNA is
degraded
OR
incorporated into chromosome by Recombination