The Polymerase Chain Reaction (PCR)

Taq polymerase

Thermas aquaticus

Producing identical copies of

a DNA segment

(typically smaller than a gene)

Manipulation of DNAMolecular Biology Techniques

Restriction enzymes: hydrolytic enzymes that

recognize a unique nucleotide sequence and

breaks (cuts) the sugar-phosphate backbone of

each DNA strand.

Single Nucleotide Polymorphisms (SNPs)

One nucleotide difference in a DNA sequence among individuals

G G C G G G C A C T

G G C G G C C A C T

Can be used for genotyping

DNA fingerprinting – Applied use

Restriction mapping whole DNA samples can be used as a “fingerprint”

This was the first approach used for DNA fingerprinting

STR site 1 STR site 2

Crime scene

DNA

AGAT GATA

The number of short tandem

repeats match

The number of short tandem

repeats do not match

GATAAGAT

Suspect’s

DNA

DNA profiling using STRs

STR = Short Tandem Repeat

4

STR = short tandem repeat

STRs are 4 nucleotide sequences

(e.g. GATA) that are repeated many times.

The number of repeats at each STR site

is highly variable among the

human population.

In forensic analysis,

STRs are amplified by PCR

5

6

Determining a person’s STR genotype at

multiple locations in the genome

builds a DNA fingerprint.

7

CODIS –Combined DNA Index System

Who is in the database?

Sex offenders, convicted felons, and in many states –

anyone arrested for a felony and in some misdeamenors

8

PTC Experiment

ampR

Aequorea victoria

• aproximately 5,400 bp

Transformation experiment

Gene CloningCLONING -- Producing identical copies of a DNA segment (gene)

Figure 20.2