5. Applications of PCR

Question 1

applications of PCR

Answer 1

diagnostics
DNA fingerprinting

Question 2

Diagnostics is used in

Answer 2

in medicine, plant pathology, environmental studies (source/tracking)

E.g. Avian sexing, genetic diseases (sickle cell trait,
cystic fibrosis, hemochromatosis, galactosaemia etc.

Question 3

DNA fingerprinting is used in

Answer 3

Identifying Individuals (Paternity Testing, Forensics)

Identifying Populations/Population Studies

Identifying Species

Question 4

DIAGNOSTICS: IDENTIFICATION OF PATHOGENIC
E. COLI STRAINS

Answer 4

Several strains of E. coli cause diarrhea and it is important to
differentiate them from non-pathogenic flora

Multiplex PCR can be used to differentiate among pathogenic
strains and non pathogenic strains

Question 5

WHAT ARE DNA FINGERPRINTS?

Answer 5

Molecular (DNA) markers that can be used to identify individuals, populations or species

Question 5

Perfect DNA Fingerprinting System

Answer 5

Large number of variable, genome wide markers

Reliable and highly polymorphic

Easy to generate & interpret

Question 6

A DNA polymorphism is

Answer 6

a sequence difference compared to a reference standard that is present in at least 1–2% of a population

Polymorphic = a gene with more than one allele

Monomorphic = gene with only one allele

Polymorphisms may or may not have phenotypic effects

Question 7

Polymorphic

Answer 7

A gene with more than one allele

Question 8

Monomorphic

Answer 8

gene with only one allele

Question 9

Polymorphisms may or may not have

Answer 9

phenotypic effects

Question 10

To identify individuals markers should be

Answer 10

polymorphic and
hypervariable

Question 11

hypervariable

Answer 11

should be unique in each individual
Recall the human genome has a lot of tandemly repeated DNA or satellite sequences

Question 12

2 main types of satellite sequences:

Answer 12

Microsatellites or STRs or SSRs – up to 13bp long

Minisatellites or VNTR – 10-100 bp repeats but hypervariable
VNTRs have repeats – 10-25 bp

Question 13

Microsatellites are usually the markers of choice to identify
individuals as

Answer 13

they are numerous (over 10,000 in the human
genome) and they are hypervariable

Question 14

STR are

Answer 14

STR are repeats of nucleotide sequences
Different alleles contain different numbers of
repeats

Question 15

STR genotypes are analyzed using

Answer 15

gel or capillary gel electrophoresis

Question 16

The FBI has chosen a set of

Answer 16

13 STR for human identity applications

PLUS Amel, an additional STR used to determine sex

Question 17

STRS – PATERNITY TESTING

Answer 17

15-20 STRs used usually 16

Each STR is named & is highly polymorphic

Each polymorphism (amplicon) is numbered

Question 18

An alleged father can be excluded as the biological father with as little as few as

Answer 18

two to three mismatches

Question 19

PATERNITY TEST — INDEX AND PROBABILITY

Answer 19

Each match receives a Paternity Index (PI) value.

The paternity index measures the strength of a particular match based on the relative uniqueness of the match

The calculation of this PI is dependent on the ethnicity of the alleged father as it is determined using allelic frequencies obtained from population data.

High PI = rare

Low PI = relatively common in population

Question 20

For an alleged father to be the child’s father the CPI value must be

Answer 20

at least 100

A CPI value of 100 corresponds to a probability of paternity of 99.0000%

A CPI value of 10,000 translates to a probability of 99.9900% and so on