Bacterial recombination and gene map or linkage map in E.coli

1. Bacterial recombination and mapping
the E-coli chromosome
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2. What is bacterial recombination?
• Bacterial recombination is a type of genetic recombination in
bacteria characterized by DNA transfer from one organism
called donor to another organism as recipient. This process
occur in three ways:
Transduction
Transformation
Conjugation
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3. Bacterial Growth
• prototrophs:
synthesize all
essential nutrients
• Auxotrophs: require a
supplement
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4. E. coli mapping
and recombination
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5. What is linkage map?
• Morgan’s student Alfred Sturtevant developed the first genetic map,
also called a linkage map.
• A linkage map is the chromosome map of a species that shows
the position of its known genes or markers relative to each
other, rather than as specific physical points on each
chromosome.
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6. Linkage Mapping
• A linkage map is a genetic map based on recombination frequencies.
• Units are called map units and shows the distance between genes.
• 1 map unit= a 1% chance of recombination.
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7. Conjugation
• Bactria undergo
conjugation , in which
• Genetic information from one
bacterium is transferred to another.
• It recombines with the second
bacterium’s DNA
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8. Conjugation in bacteria
The discovery of F+ and F- strain
• Studies of bacterial recombination began in 1946, when Joshua
Lederberg and Edward Tatum showed that bacteria undergo
conjugation, a process by which genetic information from one
bacterium is transferred to and recombined with that of another
bacterium.
• Their initial experiments were performed with two multiple
auxotrophs (nutritional mutant) of E.coli strain K12.A required
methionine(met) and biotin(bio) in order to grow, whereas strain
B required threonine(thr) , leucine(leu) and thiamine(thi).
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9. • Neither strain would grow on minimal medium. The two
strains were first grown separately in supplemented media,
and then cells from both were mixed and grown together
for several more generations.
• They were then plated on minimal medium. Any cells that
grew on minimal medium were prototrophs. It is highly
impermeable that any of the cells containing two or three
mutant gene would undergo spontaneous mutation
simultaneously at two or three independent locations to
become wild-type cells. Researchers assumed that any
prototrophs recovered must have arisen as a result of genetic
exchange and recombination between the two mutant strains.
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• Lederberg and Tatum’s elucidated the physical nature and the
genetic basis of conjugation. When the cells serve as donors
of parts of their chromosome, they are designated as F+ cells
(F for fertility). Recipient bacteria, designated as F- cells,
receive the donor chromosome material and recombine it
with part of their own chromosome.

12. Cell to cells contact is essential for chromosome
transfer
• Bernard Davis designated the Davis U-tube for growing F+ and F-
cells. When Davis plated samples from both sides of the tube on
minimal medium, no prototrophs were found and he concluded
that physical contact between cells of the two strains is essential
to gene recombination.
• Initial stage of the process of conjugation is mediated by a
structure called the F pilus (or sex pilus ) a 6-9 nm tubular
extension of the cell. Bacteria often have many pili of different
types performing different cellular functions, but all pili are
involved in some way with adhesion(the binding together of
cells).
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14. • F+ cells contain a fertility factor( F factor) that confers the
ability to donate part of their chromosome during
conjugation.
• If the infertile cells are grown with the fertile donor cells , the
F factor is regained.
Strain A Strain B
F+ F-
X
(DONOR) (RECIPIENT)
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16. Hfr Bacteria and chromosome Mapping
• In 1950, Cavalli-Sforza treated an F+ strain of E . Coli K12 with
nitrogen mustard a potent chemical known to induce
mutations. From these treated cells, he recovered a genetically
altered strain of donor bacteria that underwent recombination
at a rate of 1/104, 1000 times more frequently than the original
F+ strains.
• In 1953, William Hayes isolated another strain that demonstrated
a similarly elevated frequency of recombination. Both strains
were designated Hfr, for high-frequency recombination. Hfr cells
constitute a special class of F+ cells.
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• Another important difference was noted between Hfr strains
and the original F+ strain. If a donor cell is from an Hfr
strain , recipient cells, though sometimes displaying genetic
recombination, never become Hfr, thus they remain F-.
• In comparison then,
• The most significant characteristics of Hfr strains is the
specific nature of recombination in each case. In a given
Hfr strain, certain genes are more frequently recombined
than others, and some do not recombine at all.

18. • In the mid- 1950s , experiment by Ellie Wollman and
Francois Jacob explained the differences between Hfr cells
and F+ cells and showed how Hfr strains would allow
genetic mapping of the E .coli chromosome.
• A culture containing a mixture of an Hfr and F- strain was
incubated , and samples were removed at intervals and
placed in a blender. The shear forces created in a blender
separated conjugating bacteria so that the transfer of the
chromosome was determined. Cells are subsequently tested
for the transfer of specific genes. This process is called the
interrupted mating technique.
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19. conjugation
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20. • Thirty-four hundred lambda phage clones containing
segments of the E. coli chromosome were isolated and used
to construct a 4700 kb long integrated restriction map for
eight six-base-recognizing enzymes by a rapid mass-analysis
method.
• This technique was utilized to map all genes of E. coli
chromosome.
• 100 minutes long (how long it takes to transfer over the
entire chromosome)
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21. • During the first 8 minutes after the two strains were mixed,
no genetic recombination was detected. At about 10
minutes , recombination of the aziR gene could be detected.
By 15 minutes 50% of recombinants were aziR and 15%
were also tonS. Within 20 minutes lat+ gene was found
among the recombinants and within 25 minutes gal+ was
also beginning to transferred.
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23. conclusion
• An ordered transfer of gene that correlated with the
length of time conjugation proceeded. Gene order and
distance between genes, as measured in minutes, could be
predicted from experiments.
• This information sometimes referred to as time mapping,
served as the basis for the first genetic map of the E.coli
chromosome. Minutes provide a measure similar to map
units.
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24. Types of gene mapping
 Ganetic mapping: interlocus distance
 linear description of markers and genes on a
given chromosome with markers closer being inherited
together more often.
 Physical mapping: location of genes on chromosome
 cytogenetics
 physical methods
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26. Gene Mapping
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27. Transformation
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• In transformation, small
pieces of extracellular
DNA are taken up by
a living bacterial cell
and integrated stably
into the chromosome.

28. Transduction
• Type of bacterial genetic recombination caused by the
infection of a bacteriophage
• Bacteriophage can infect a host bacterium by injecting
their DNA.
• Lysogeny occurs when:
the phage DNA integrates into the bacterial chromosome
it is replicated along with the chromosome
it is passed to daughter cells
• Bacteria containing a prophage are lysogenic and can grow
and divide stably until viral reproduction is induced.
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• In generalized transduction, bacterial DNA instead of
phage DNA is packaged in a phage particle and
transferred to a recipient host.
• In specialized transduction, a small piece of
bacterial DNA is packaged along with the phage
DNA.

30. Transduction : Generalized
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31. Transduction : Specialized
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