Fig. 12-00
Fig. 12-01
Fig. 12-02
Fig. 12-03
Fig. 12-04
Fig. 12-05
Fig. 12-06
Fig. 12-07
Plasmids
Bacterialchromosome
Remnant ofbacterium
Co
lori
zed
TE
M
Fig. 12-08-1
Plasmid
Bacterial cell
Isolateplasmids.
Fig. 12-08-2
Plasmid
Bacterial cell
Isolateplasmids.
DNA
IsolateDNA.
Cell containingthe gene of interest
Fig. 12-08-3
Plasmid
Bacterial cell
Isolateplasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Cell containingthe gene of interest
Fig. 12-08-4
Plasmid
Bacterial cell
Isolateplasmids.
Gene of interest
Recombinant DNA plasmids
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Fig. 12-08-5
Plasmid
Bacterial cell
Isolateplasmids.
Recombinant bacteria
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Fig. 12-08-6
Plasmid
Bacterial cell
Isolateplasmids.
Clone the bacteria.
Recombinant bacteriaBacterial clone
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Fig. 12-08-7
Plasmid
Bacterial cell
Isolateplasmids.
Find the clone withthe gene of interest.
Clone the bacteria.
Recombinant bacteriaBacterial clone
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Fig. 12-08-8
Plasmid
Bacterial cell
Isolateplasmids.
Some usesof genes
Gene for pestresistance
Gene fortoxic-cleanupbacteria
Genes may beinserted intoother organisms.
Find the clone withthe gene of interest.
The gene and proteinof interest are isolatedfrom the bacteria.
Clone the bacteria.
Recombinant bacteriaBacterial clone
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
Harvestedproteins may beused directly.
Some usesof proteins
Protein for“stone-washing”jeans
DNA
Cell containingthe gene of interest
Protein fordissolvingclots
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Fig. 12-09-1 Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
A restriction enzyme cutsthe DNA into fragments.
Fig. 12-09-2 Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
A DNA fragment is addedfrom another source.
A restriction enzyme cutsthe DNA into fragments.
Fig. 12-09-3 Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
A DNA fragment is addedfrom another source.
A restriction enzyme cutsthe DNA into fragments.
Fragments stick together bybase pairing.
Fig. 12-09-4 Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
DNAligase
Recombinant DNA molecule
A DNA fragment is addedfrom another source.
A restriction enzyme cutsthe DNA into fragments.
Fragments stick together bybase pairing.
DNA ligase joins thefragments into strands.
Fig. 12-10
Radioactive probe(single-stranded DNA)
Single-stranded DNA
Mix with single-stranded DNA fromvarious bacterial clones
Base pairing indicates thegene of interest
Fig. 12-11-1Cell nucleus
DNA ofeukaryoticgene
Test tube
Transcription
Exon Intron Exon ExonIntron
Fig. 12-11-2Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
Transcription
Introns removed andexons spliced together
Exon Intron Exon ExonIntron
Fig. 12-11-3Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
Reversetranscriptase
Transcription
Introns removed andexons spliced together
Isolation of mRNA fromcell and addition ofreverse transcriptase
Exon Intron Exon ExonIntron
Fig. 12-11-4Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
Reversetranscriptase
cDNA strandbeing synthesized
Transcription
Introns removed andexons spliced together
Isolation of mRNA fromcell and addition ofreverse transcriptase
Synthesis of cDNAstrand
Exon Intron Exon ExonIntron
Fig. 12-11-5Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
cDNA of genewithout introns
Reversetranscriptase
cDNA strandbeing synthesized
Transcription
Introns removed andexons spliced together
Isolation of mRNA fromcell and addition ofreverse transcriptase
Synthesis of cDNAstrand
Synthesis of second DNAstrand by DNA polymerase
Exon Intron Exon ExonIntron
Fig. 12-12
Fig. 12-13-1
DNA isolated
Crime scene Suspect 1 Suspect 2
Fig. 12-13-2
DNA isolated
DNA amplified
Crime scene Suspect 1 Suspect 2
Fig. 12-13-3
DNA isolated
DNA amplified
DNA compared
Crime scene Suspect 1 Suspect 2
Fig. 12-14
Fig. 12-14a
Fig. 12-14b
Fig. 12-15
InitialDNAsegment
Number of DNA molecules
1 2 4 8
Fig. 12-16
Crime scene DNA
Suspect’s DNA
Same number ofshort tandem repeats
Different numbers ofshort tandem repeats
STR site 1 STR site 2
AGAT
AGAT GATA
GATA
Fig. 12-17-1
Mixture of DNAfragments ofdifferent sizes
Powersource
Gel
Fig. 12-17-2
Mixture of DNAfragments ofdifferent sizes
Powersource
Gel
Fig. 12-17-3
Mixture of DNAfragments ofdifferent sizes
Powersource
Gel
Completed gel
Band oflongest(slowest)fragments
Band ofshortest(fastest)fragments
Fig. 12-18
Amplifiedcrime sceneDNA
Amplifiedsuspect’sDNA
Longerfragments
Shorterfragments
Fig. 12-19
Crime sceneDNA
Suspect’sDNA
Fragment w
Fragment x
Fragment y
Longerfragments
Shorterfragments
Fragment z
Fragment y
Crime sceneDNA
Suspect’sDNA
Cut
Cut Cut
Restriction enzymes added
x
wy y
z
Fig. 12-20
Fig. 12-21
Anthraxspore
Envelopecontaininganthrax spores
Fig. 12-22-1
Chromosome
Fig. 12-22-2
Chromosome
Chop up withrestriction enzyme
DNA fragments
Fig. 12-22-3
Chromosome
Chop up withrestriction enzyme
Sequencefragments
DNA fragments
Fig. 12-22-4
Chromosome
Chop up withrestriction enzyme
Sequencefragments
DNA fragments
Alignfragments
Fig. 12-22-5
Chromosome
Chop up withrestriction enzyme
Sequencefragments
DNA fragments
Alignfragments
Reassemblefull sequence
Fig. 12-22a
Fig. 12-23
Fig. 12-24-1
Normal humangene isolatedand cloned
Healthy person
Fig. 12-24-2
Normal humangene isolatedand cloned
Normal humangene insertedinto virus
Healthy person
Harmlessvirus (vector)
Virus containingnormal human gene
Fig. 12-24-3
Normal humangene isolatedand cloned
Normal humangene insertedinto virus
Virus injectedinto patient withabnormal gene
Healthy person
Harmlessvirus (vector)
Virus containingnormal human gene
Bonemarrow
Bone of personwith disease
Fig. 12-25
Fig. 12-26
Fig. 12-27
Fig. 12-28
Fig. 12-T01
Fig. 12-UN01
DNA isolated from twosources and cut by samerestriction enzyme
Gene of interest(could be obtained froma library or synthesized)
RecombinantDNA
Plasmid(vector)
Transgenic organisms
Useful products
Fig. 12-UN02
Crime scene Suspect 1 Suspect 2
DNA
Polymerase chainreaction (PCR)amplifies STRsites
LongerDNAfragments
ShorterDNAfragments
DNA fragments compared by gel electrophoresis
Gel
Fig. 12-UN03
Normalhuman gene
Virus
Bonemarrow
Normal human gene is transcribedand translated in patient, potentiallycuring genetic disease permanently