e2_p2 Flashcards
Co-transformation of linked genes
genes located close together are often transferred as a unit to recipient cell = co-transformation
co-transformation of 2 genes at frequency greater than product of the single-gene transformation frequencies implies the 2 genes are located close together Genes that are far apart are less likely to be transferred together.
Co-transformation is used to map gene order
Hfr cell
Cells with the F plasmid integrated into the bacterial chromosome
When an Hfr cell undergoes conjugation, the process of transfer of F factor is initiated in the same manner as in an F+ cell
However, because F factor is part of the bacterial chromosome transfer from an Hfr cell also includes DNA from the bacterial chromosome
F factor & conjugation
F (fertility) factor is a conjugative plasmid transferred from cell to cell by conjugation.
It is an episome.
The F plasmid is a low-copy-number plasmid (1-2 copies per cell) and ~100 kb in length
It replicates once per cell cycle and segregates to both daughter cells in cell division.
E.Coli as a model organism
1 circular DNA molecule
4.6 million bp
haploid
divides once every 20-60 min (depending on culture media)
Abnormalities of Structure : Insertion
Non-Reciprocal translocation
Segment from one chromosome is inserted into a different chromosome - requires 3 chromosome breaks (rare)
A chromosomal duplication is usually caused by
abnormal events during recombination
Telomerase activity
different level of activity in different cells
no activity in most somatic cells
Moderately repetitive
Found a few hundred to few thousand times
Genes for rRNA & histones
Sequences that regulate gene expression and translation.
Transposable elements.
Unique (or non-repetitive sequences)
Found once or a few times in the genome.
Includes structural genes as well as intergenic areas.
In humans, makes up roughly 41% o the genome.
Highly repetitive
Found 10s of 1000s to millions of times
Each copy is relatively short (a few nucleotides to several hundred in length)
Some sequences are interspersed throughout genome.
-Example : Alu family in humans
Other sequences are clustered together in tandem arrays -> these are commonly found in the centromeric regions
Alu family in humans
highly repetitive sequences
Approximately 300 bp long
Represents 10% of the human genome
Found every 5000-6000 bp
3 implications of Hfr cell conjugation
- Recipient usually does not receive the entire chromosome, nor F plasmid (so remains F-)
- Genes proximal to 5’ end of F integration site transferred most frequently.
- Homologous recombination between ‘donated’ DNA & recipient. If donor is wild & recipient is mutant, DNA donation can be tested.
If donor is wild-type and recipient is mutant, DNA donation can be tested (how?)
We can map the relative position of genes versus F integration site.
Genes close to integrated F factor are always transferred, but the further away from F position, the less likely.
How many minutes does mixing of Hfr donor and F- recipient need to last until a certain gene is transferred?
->Every gene has unique ‘minute’ location on genome, (complete genome transfer 100 min)
Mapping Problem
The distance between genes is determined by comparing their times of entry during an interrupted mating experiment.
The approximate time of entry is computed by extrapolating the time back to the x-axis.
Interrupted Mating Experiment
Mix togetehr Hfr donor cells and F- cells
After different periods of time, take some and place in a blender (breaks about the cells).
Plate on growth medium to detect gene transfer.
