campbell dna and technology

BIOLOGYCONCEPTS & CONNECTIONS

Fourth Edition

Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings

Neil A. Campbell • Jane B. Reece • Lawrence G. Mitchell • Martha R. Taylor

From PowerPoint® Lectures for Biology: Concepts & Connections

CHAPTER 12DNA Technology and the Human Genome

Modules 12.1 – 12.6


• Research on E. coli revealed that these bacteria have a sexual mechanism that can bring about the combining of genes from two different cells

• This discovery led to the development of recombinant DNA technology

– a set of techniques for combining genes from different sources

From E.Coli to a Map of Our Genes


• DNA technology has many useful applications– The Human Genome Project

– The production of vaccines, cancer drugs, and pesticides

– Engineered bacteria that can clean up toxic wastes


• Transformation, the taking up of DNA from the fluid surrounding the cell

12.1 In nature, bacteria can transfer DNA in three ways

BACTERIA AS TOOLS FOR MANIPULATING DNA

Figure 12.1A

DNA enterscell

Fragment ofDNA from anotherbacterial cell

Bacterial chromosome(DNA)


• Transduction, the transfer of bacterial genes by a phage

• Conjugation, the union of cells and the DNA transfer between them

Fragment ofDNA from anotherbacterial cell(former phagehost)

Phage

Sex pili

Mating bridge

Donor cell(“male”)

Recipient cell(“female”)

Figure 12.1CFigure 12.1B


• The transferred DNA is then integrated into the recipient cell’s chromosome

Figure 12.1D

Donated DNA

Recipient cell’schromosome

Crossovers

Degraded DNA

Recombinantchromosome


• An F factor is a DNA segment in bacteria that enables conjugation and contains an origin of replication

12.2 Bacterial plasmids can serve as carriers for gene transfer

Figure 12.2A

F factor (integrated)

Male (donor) cell

Origin of F replication

Bacterial chromosome

F factor startsreplication andtransfer of chromosome

Only part of thechromosome transfers

Recipient cell

Recombination can occur


• An F factor can exist as a plasmid, a small circular DNA molecule separate from the bacterial chromosome

Figure 12.2B, C

F factor (plasmid)

Male (donor) cell

Bacterial chromosome

F factor startsreplication andtransfer

Plasmid completestransfer andcircularizes

Cell now male

Plasmids


• Plasmids are key tools for DNA technology

– Researchers use plasmids to insert genes into bacteria

12.3 Plasmids are used to customize bacteria: An overview


Figure 12.3

Plasmidisolated

1Bacterium

Bacterialchromosome

Plasmid

2DNAisolated

Cell containing geneof interest

DNAGene ofinterest

3 Gene inserted into plasmid

Recombinant DNA(plasmid)

4 Plasmid put intobacterial cell

Recombinantbacterium

5

Copies of gene Copies of protein

Clones of cellGene for pestresistanceinserted intoplants

Gene used to alter bacteriafor cleaning up toxic waste

Protein used to dissolve bloodclots in heart attack therapy

Protein used to make snow format highertemperature

Cell multiplies withgene of interest


• Restriction enzymes cut DNA at specific points

• DNA ligase “pastes” the DNA fragments together

• The result is recombinant DNA

12.4 Enzymes are used to “cut and paste” DNA

Figure 12.4

DNA1

Restriction enzymerecognition sequence

Restriction enzymecuts the DNA intofragments

Sticky end

2

3

4

5

Restriction enzymecuts the DNA intofragments

Addition of a DNAfragment fromanother source

Two (or more)fragments sticktogether bybase-pairing

DNA ligasepastes the strand

Recombinant DNA molecule


• Bacteria take the recombinant plasmids and reproduce

• This clones the plasmids and the genes they carry

– Products of the gene can then be harvested

12.5 Genes can be cloned in recombinant plasmids: A closer look


Figure 12.5

Isolate DNAfrom two sources

1E. coli

Cut both DNAs with the same

restrictionenzyme

2

Plasmid

Human cell

DNA

Gene VSticky ends

Mix the DNAs; they joinby base-pairing

3

Add DNA ligaseto bond the DNA covalently

4

Recombinant DNAplasmid Gene V

Put plasmid into bacteriumby transformation

5

Clone the bacterium6

Bacterial clone carrying manycopies of the human gene


• Recombinant DNA technology allows the construction of genomic libraries

– Genomic libraries are sets of DNA fragments containing all of an organism’s genes

• Copies of DNA fragments can be stored in a cloned bacterial plasmid or phage

12.6 Cloned genes can be stored in genomic libraries

Figure 12.6

Genome cut up with restriction enzyme

Recombinantplasmid

OR

Recombinantphage DNA

Bacterialclone

Phageclone

Phagelibrary

Plasmidlibrary

campbell dna and technology

Documents