DNA Technology

Biotechnology

The use or alteration of cells or biological molecules for specific applicationsTransgenics

Transgenic“changed genes”

Recombinant DNADNA from different species mixed together

Natural or man-madeWhole organisms or cellsPossible because the genetic code is universal

All life uses the same genetic code (A,T,G,C)

Amplifying DNA

Need many copies for various DNA tests from a small initial sample

Two techniquesPolymerase chain reaction (PCR)

Recombinant DNA technology

PCR

Done on moleculesBased on DNA replicationRapidly replicates a selected sequence of DNA in a test tubeUsed to:

Establish blood relationshipsIdentify remainsConvict or exonerate suspectsLook at pathogensIdentify genes

Very sensitive but easily contaminated

Review of DNA Replication

Parentalmoleculeof DNA

Both parentalstrands serveas templates

Two identicaldaughter molecules

of DNA

T

Nucleotides

C G

A

G C

A

A

T

T

A

AA

A

C C

C

T

T

G

G

A

C

A

A A

A

A

A

C C

C

CG

T

G

T

T

T

T

G

T

T

T

G

G

Requirements for PCR

Know parts of the DNA sequence to be amplified2 DNA Primers

Short, lab-made single-stranded DNAOne complementary for each strand in the DNA segment to be amplified

Lots of nucleotidesTaq DNA polymerase

From Thermus aquaticus (lives in hotsprings)

A thermal cycler

PCR

Recombinant DNA (cloning)

Works in cells

Adds genes from one type of organism to the genome of another

Requires:Restriction enzyme

Vector

Donor DNA

Host bacteria

Restriction Enzymes

Figure 19.3a

Cloning Vectors

Carries DNA from the cells of one species into cells of another

Any piece of DNA into which another can be inserted

Types of Vectors

Which vector used depends on size of the gene to be inserted

PlasmidOne type of cloning vector

Small circle of double-stranded DNA

Occurs naturally in some bacteria and yeasts

Creating Recombinant DNA Molecules

Figure 19.3a

Figure 19.3b

Recombinant DNA

Figure 19.5

Selecting Recombinant DNA Molecules

3 types of cellsCells that lack plasmidsCells that contain plasmids that do not contain a foreign geneCells that contain plasmids that have the foreign gene –WANT THESE

Plasmids can contain a gene for an enzyme that catalyzes a reaction that makes a blue color

If a foreign gene inserts in the middle of this “blue” gene, the bacteria containing the plasma will not produce the blue color

Plasmids can contain a gene for antibiotic resistance

When the antibiotic is applied to the bacterial cells, only those with the plasmid will survive

Applications of Recombinant DNA

Drugs (e.g. insulin)Pure, human versions

e.g. “humulin”

Transgenic plantsa.k.a. genetically modified (GM) plants

Transgenic animalse.g. 3xTg-AD mouse

Transgenic Plants

Transgenic Animals

Isolating a Gene

Make a DNA probeStrand of DNA complementary to the one of interest

Attached to a radioactive or fluorescent molecule

Complementary (cDNA) Library

Represents only protein-encoding genes

Made from mRNAsRepresents the proteins being made in a particular cell

Making a cDNA library

Extract mRNAs from cells

Use reverse transcriptase to make DNA from RNA

Use DNA polymerase to make double-stranded DNA