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