Sept 29, 2009
Formation of a Recombinant DNA Genome
-A stretch of mammalian DNA is first sliced up by restriction enzymes to create
minor fragments for cloning. The fragments can be inserted into bacterial
fragments via hybridization (DNA-ligase base pairing) and then cloned by PCR.
-The human genome can generate around 1 million recombinant DNA molecules.
-Once the recombinant DNA molecules have been successfully hybridized onto a
bacterial plasmid it needs to be plated onto a gel for growth. The hybrids are
sparsely placed onto a gel and then the desired gene can be selected for.
-Transfer of DNA recombinants is usually inefficient and only a small portion of
the population will accept the hybridized gene.
***refer to previous notes for screening process*** Lambda (λ) Phage
-The phage infects an E. coli and begins a lytic cycle; the process follows a series
lytic cycle replicates on its on lysis E. coli to release progeny.
-The lysogenic cycle allows the phage to become dormant and emerges later on;
there is also series of steps that it follows:
lysogenic cycle integrates into E. coli genome (prophage) replicates along
with the E. coli genome
-These phage can revert between the two cycles depending on the circumstances.
In order for the phage to select between the lysogenic or lytic cycle it just splices
itself into different regions of the bacterial genome
-Refer to fig. 6.22 Singer and Burk
-this process is much faster than picking colonies since the nitrocellulose can be
applied directly onto the plates.