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Genetic Frontiers. Chapter 15. LEARNING OBJECTIVE 1. Define genetic engineering Outline the primary techniques used in recombinant DNA technology , including genetic probes and DNA cloning. KEY TERMS. GENETIC ENGINEERING Manipulation of genes, often through recombinant DNA technology

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learning objective 1
LEARNING OBJECTIVE 1
  • Define genetic engineering
  • Outline the primary techniques used in recombinant DNA technology, including genetic probes and DNA cloning
key terms
KEY TERMS
  • GENETIC ENGINEERING
    • Manipulation of genes, often through recombinant DNA technology
  • RECOMBINANT DNA TECHNOLOGY
    • Techniques used to make DNA molecules by combining genes from different organisms
recombinant dna technology 1
Recombinant DNA Technology 1
  • DNA molecules are cleaved at specific base sequences
    • to break them into smaller fragments
  • Segments of DNA from different sources are joined, forming recombinant DNA molecule
    • these molecules are taken into a host cell
recombinant dna technology 2
Recombinant DNA Technology 2
  • Cells that take up the gene are identified with a genetic probe
  • The gene may be transcribed and translated within the host cell
    • leading to production of a protein not previously produced by the host organism
key terms1
KEY TERMS
  • GENETIC PROBE
    • A single-stranded nucleic acid used to identify a complementary sequence by base pairing with it
slide9

Bacterial

colonies

Filter paper

1

Bacterial cells are spread on solid

nutrient medium so that only one cell

is found in each location. Each cell

multiplies to give rise to genetically

identical descendants that form a

colony on the medium. A few cells

from each colony are transferred to

special filters.

1

Filter with bacteria from colonies; cells are broken to

expose the DNA

Radioactively

labeled nucleic acid probe is added

2

Radioactively labeled probe nucleic

acid is added to the filter; the probe

nucleic acid is single stranded and

contains a sequence of nucleotides

complementary to the gene of interest.

2

3

Some radioactive

nucleic acid probe

molecules form base pairs with the DNA of some colonies

3

Some of the probe nucleic acid forms

base pairs with the DNA of some of

the colonies.

Exposed X-ray film; dark spots identify colonies with desired DNA

4

DNA from bacterial cells that contain

the DNA sequence complementary to

the radioactive probe can be detected

by X-ray film.

4

Fig. 15-6, p. 299

key terms2
KEY TERMS
  • DNA CLONING
    • Process of selectively amplifying DNA sequences so their structure and function can be studied
learning objective 2
LEARNING OBJECTIVE 2
  • Explain the actions and importance of restriction enzymes and ligase
key terms3
KEY TERMS
  • RESTRICTION ENZYME
    • An enzyme used in recombinant DNA technology to cleave DNA at specific base sequences
    • Breaks the DNA molecule into more manageable fragments
ligase
Ligase
  • Segments of DNA from different sources are joined by the enzyme ligase
slide15

Site of cleavage

C

T

T

G

A

A

A

A

G

T

T

C

Site of cleavage

Sticky end

C

T

A

A

G

T

A

G

T

C

A

T

Sticky end

Fig. 15-2, p. 295

learning objective 3
LEARNING OBJECTIVE 3
  • Identify the role of biological vectors, such as plasmids, in recombinant DNA technology
  • Describe a biological vector and a nonbiological method used to introduce genes into plant cells
key terms4
KEY TERMS
  • VECTOR
    • An agent, such as a plasmid or virus, that transfers DNA from one organism to another
  • PLASMID
    • A small, circular DNA molecule that carries genes separate from the main DNA of a bacterial cell
slide19

Main bacteria DNA

Bacterium

Plasmid

Fig. 15-3, p. 296

a biological vector
A Biological Vector
  • The plasmid of Agrobacterium is an effective vector for introducing genes into many plant cells
slide22

DNA from

another

organism

Sticky

end

Sticky

end

. . . treated with a

restriction enzyme

Plasmid from a

bacterium . . .

Treated

with the same

restriction enzyme

A plasmid and

plant DNA are

spliced together

with DNA ligase

Fig. 15-4, p. 297

slide23

DNA from

another

organism

Sticky

end

Sticky

end

. . . treated with a

restriction enzyme

Plasmid from a

bacterium . . .

Treated

with the same

restriction enzyme

A plasmid and

plant DNA are

spliced together

with DNA ligase

Stepped Art

Fig. 15-4, p. 297

slide25

Sites of cleavage

Fragment

2

Fragment

3

Fragment

1

Fragment

4

Plant DNA

Cut with a restriction enzyme

1

1

DNA from plant cells is cut into multiple fragments with a restriction enzyme. (Only a small part of one chromosome is shown.)

Produce recombinant

DNA

2

2

2

2

Recombinant plasmids are formed by cutting plasmids with the same restriction enzyme, mixing the plasmids with the segments of plant DNA, and treating with ligase.

2

Gene for

resistance

to antibiotic

3

3

Because the recombinant plasmids retain a gene for resistance to an antibiotic (R), bacterial cells that contain the plasmids are resistant to that antibiotic.

Plate with antibiotic-containing

medium

The bacteria are then grown on an antibiotic-containing nutrient medium, and only those that contain the recombinant plasmid survive.

4

4

Bacteria without

plasmid fail to grow

Bacteria with plasmid

live and multiply

Fig. 15-5, p. 298

a nonbiological method
A Nonbiological Method
  • A nonbiological approach to introduce DNA into plant cells is a genetic “shotgun”
  • Researchers coat microscopic gold or tungsten fragments with DNA and then shoot them into plant cells
slide28

Plasmid

Foreign gene

Antibiotic-resistant gene

Genetically

engineered

plant cells

Cultured

plant cells

Pieces of plant tissue are placed in a suitable medium, and the cells grow to form a clump of undifferentiated cells.

1

Foreign DNA is spliced into the crown gall plasmid. The recombinant plasmid is

inserted into Agrobacterium

tumefaciens, which infects plant cells in culture.

The plant cells divide

in tissue culture. Each cultured plant cell contains the foreign gene.

Genetically engineered plants are produced from the cultured plant

cells through the use of plant tissue culture techniques.

2

3

4

Fig. 15-8, p. 301

learning objective 4
LEARNING OBJECTIVE 4
  • Define DNA sequencing
key terms5
KEY TERMS
  • DNA SEQUENCING
    • Procedure by which the sequence of nucleotides in DNA is determined
dna sequencing
DNA Sequencing
  • Automated DNA-sequencing machines connected to powerful computers let scientists sequence huge amounts of DNA quickly and reliably
learning objective 5
LEARNING OBJECTIVE 5
  • Define genome
  • Briefly describe the emerging field of genomics
key terms6
KEY TERMS
  • GENOME
    • All the genetic material contained in an individual
key terms7
KEY TERMS
  • GENOMICS
    • Field of biology that studies the genomes of various organisms
    • Tries to identify all the genes, determine their RNA or protein products, and ascertain how the genes are regulated
learning objective 6
LEARNING OBJECTIVE 6
  • Explain how RNA interference is used to study gene function
key terms8
KEY TERMS
  • RNA INTERFERENCE(RNAi)
    • Makes use of certain small RNA molecules that interfere with the expression of genes or their RNA transcripts
rna interference rnai
RNA Interference (RNAi)
  • After a protein-coding gene is identified, the function of that gene is studied using RNAi to shut the gene off
  • After the gene is silenced, biologists observe changes in phenotype to determine function of missing protein
learning objective 7
LEARNING OBJECTIVE 7
  • Describe at least one application of recombinant DNA technology in each of the following: medicine and pharmacology, DNA fingerprinting, and transgenic organisms, specifically genetically modified crops
medicine and pharmacology
Medicine and Pharmacology
  • Escherichia coli have been genetically engineered to produce human insulin
    • Significant medical benefits to insulin-dependent diabetics
dna fingerprinting
DNA Fingerprinting
  • Analysis of DNA from an individual
  • Applications
    • Investigating crime (forensic analysis)
    • Studying endangered species in conservation biology
    • Clarifying disputed parentage
key terms9
KEY TERMS
  • TRANSGENIC ORGANISM
    • A plant or other organism that has foreign DNA incorporated into its genome
  • GENETICALLY MODIFIED (GM) CROP
    • A crop plant that has had its genes intentionally manipulated (transgenic crop plant)
genetically modified gm crops
Genetically Modified (GM) Crops
  • Agricultural geneticists developing GM plants that are resistant to insect pests, viral diseases, drought, heat, cold, herbicides, and salty or acidic soil
learning objective 8
LEARNING OBJECTIVE 8
  • Discuss safety issues associated with recombinant DNA technology
  • Explain how these issues are being addressed
safety issues 1
Safety Issues 1
  • Concerns: Genetically engineered organisms might be dangerous if they escaped into the environment
    • Scientists carried out risky experiments in facilities designed to hold pathogenic organisms
safety issues 2
Safety Issues 2
  • So far, there is no evidence that researchers have accidentally cloned hazardous genes or released dangerous organisms into the environment
    • Scientists have relaxed many of restrictive guidelines for using recombinant DNA
safety issues 3
Safety Issues 3
  • Stringent restrictions still exist where questions about possible effects on the environment are unanswered
    • (Example: in research that proposes to introduce transgenic organisms into the wild)