Escolar Documentos
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Transformation
Transformation: DNA picked up
directly from the medium and
recombined into the genome
heteroduplex
Competent cell:
capable of picking
up DNA
Transduction
~1 phage/10,000 will
pick up chromosomal
DNA...
Plasmids
Conjugation
Resistance Plasmids
Genetic Engineering
Restriction Endonucleases
• A Restriction Endonucleases will cut both
strands of a DNA duplex at a specific place
• These “places” need not be directly opposite:
5’…GAATTC…3’
3’…CTTAAG…5’
5’…G -OH5’…GAATTC…3’
P-AATTC…3’
3’…CTTAA3’…CTTAAG…5’
-P HO-G…5’
G^AATT-C G^GATC-C
C-TTAA^G C-CTAG^G
Nodeba Bob Abedon
A^GATC-C GC^GGCC-GC
T-CTAG^G CG-CCGG^CG
Complementary (c)DNA Note that the reverse
transcriptase is primed
by the poly-A found at
the end of most
eukaryotic mRNAs
Note that there are
issues (& variation) as
to how one makes the
second DNA strand
Here that strand is
generated using a
second enzyme that is
primed via generation
of a hairpin of DNA by
reverse transcriptase
Ligation into Plasmid
Other vectors
include viruses
(transduction) as
well as
otherwise inert
projectiles
Virus (Phage) Vector
Polymerase Chain Reaction
Gel Electrophoresis (1/2)
Gel Electrophoresis (2/2)
Southern Blotting
Visualized DNA
Restriction Fragment Length Polymorphism
DNA Sequencing
DNA Microarrays
Monoclonal Antibodies (Mabs)
http://www.healthsci.utas.edu.au/hls/units/cxa262/lectures/cloning.ppt
www.healthsci.utas.edu.au/hls/units/ cxa262/lectures/cloning.ppt
www.ou.edu/cas/zoology/Courses/3333/powerpoint/lec8.ppt
http://www.ipam.ucla.edu/publications/fg2000/fgt_tspeed8.ppt
http://www.bioscience.drexel.edu/Homepage/Fall2002/Bio449/powerpoints/BIO449%20lab%20lecture%203.ppt