The molecular movement of the polymerase similar to opening and

closing of the hand:

occurs once per replication

occurs once for each nucleotide

occurs several times for each nucleotide

How can you mimic the DNA polymerase "in action"?

With your right hand.

With your left hand.

With both hands.

With either hand.

Which DNA end is represented by the upper end of the metal stick
representing the template?

5' end

3' end

Why does DNA polymerase open?

To elongate the template strand.

To allow entry of nucleotides.

To check for proper base pairing.

To catalyse the reaction.

Why does DNA polymerase close?

To allow entry of nucleotides.

To check for proper base pairing.

To catalyse the reaction.

Is it more likely that DNA polymerase is open or closed, if it has to
remove a nucleotide after wrong base pairing?

Open

Closed

The human genome has approx.:

3 million base pairs

3 billion base pairs

3 trillion base pairs

The error rate of DNA polymerase is

1: 1000

1: 1 million

1: 1 billion

Why is the error rate 6000 per replication?

Because each chromosome is present twice.

Because both strands are copied.

Because 3 Billion base pairs means 6 Billion bases.

An exonuclease can cleave nucleotides:

from the middle of a strand

from the end of a strand

The proof-reading (koreksi cetakan percobaan) function:

reduces the error rate

slows down replication

corrects about 99% of replication errors

is the reason why DNA is synthesized only in 5 to 3 direction


An error in replication:

causes a mutation

always causes disease

might cause a disease if it occurs at a sensitive site


PCR is called the polymerase chain reaction. Why is that?

Because DNA is a long chain of nucleotides.

Because the amount of product of the reaction increases in an exponential fashion.

Because 'poly' means 'many'.

Because its impact on science was like a nuclear explosion.


We're starting from one double stranded DNA molecule, with primers
that fit properly. After three rounds of PCR, how many strands of DNA
have we got?

16

3

4

8


PCR is used to:

amplify the existing amount of DNA in a test tube, without altering its sequence

produce new DNA without using a DNA template

In a PCR, the amount of product increases:

linearly

exponentially


The region/s that is/are amplified in a PCR:

are the ones that are bound by the two primers

is the one in between the two primers, including those that are bound by the two
primers

are the ones outside the two primers

is the entire DNA molecule


How many DNA molecules are present after 5 rounds of PCR on a
single DNA molecule?

16

32

64

5

What can be used as a sample for PCR?

Mucosal cells

Blood

Protein extracts

Cytoplasm

What are the advantages of PCR? It is ...

small amounts of sample can be used

fast

inexpensive

color-coded

very specific


Sickle cell anemia is caused by a point mutation, meaning:

a deletion in 1 region

the exchange of 1 base in a gene

one gene is missing

PCR is highly suitable to detect:

all types of cancer

monogenetic diseases

diseases with unknown reason


Autosomal recessive means:

the defect is on the X chromosome

a defect on only one chromosome will lead to disease

the defect is on any chromosome except X and Y

a healthy person can still be a carrier

Why is PCR very interesting when bacteria and viruses are to be
detected?

They can't be detected with other methods

Other methods might require cultivation which is risky for personnel

Cultivation required for other methods might be difficult

Which diseases can be diagnosed with PCR?

Sickle cell anemia

Chorea Huntington

Cystic fibrosis

Tuberculosis


The colorless tubes already contain lyophilized:

primers

salts

nucleotides

polymerase
The kit used in the video is a kit for detection of:

Mycoplasma

Tuberculosis

Viruses

Andrei and Can have to add:

a buffer

the samples

the polymerase

water for the negative control


The red tubes contain :

a negative control

a positive control (mycoplasma DNA)
The PCR is run for approx. 90min in:

a PCR machine

an oven

in the incubator


In the meantime, Andrei and Can prepare an agarose gel:

for sample collection

for sample separation

for sample storage


The PCR product is directly loaded onto the gel for electrophoresis
during which:

DNA is synthesized

DNA is separated due to size

negatively charged DNA migrates towards the positive pole in an electrical field

DNA becomes visible


Then, the gel is placed into an ethidium bromide solution for:

fixing

staining

development

Ethidium bromide is an intercalating dye and therefore:

is dangerous

needs to be handled with care

can cause mutations

helps to visualize DNA later

can be replaced with less dangerous GelRed if available


Upon excitation with UV light Ethidium bromide (EtBr) emits
fluorescence:

that is stronger when EtBr has intercalated

that is weaker in areas without DNA

that allows us to see the DNA bands

that is dangerous


On the final image two bands can be present in the samples and
controls:

that are of different size

that indicate mycoplasma DNA and internal control DNA

that indicate different types of mycoplasma

that indicate whether the cells are contaminated

What does STR stand for?

Small terrific repeats

Small tandem repeats

Short tandem repeats

Specific tandem repeats


Which fields have truly been revolutionized by PCR?

Forensics

Nutritional Research

Pediatrics

Crime scene investigation

How many regions are used for genetic fingerprinting by the FBI?

13

15

23

46

What is the analysis of "amelogenin" included for?

None of the above.

To show that the genetic fingerprinting has worked.

To reveal mistakes.

To determine gender.


Why are STRs so well suited for genetic fingerprinting?

They are short.

They are repetitive.

They are frequent.

They occur in different numbers.

All of the above

What does
tandem
in the term STR actually mean in terms of sequence?

short in length

identical sequence on both strands

repetitive sequence of nucleotides

a defined number of repeats

If the STR sequence on one strand reads 5'-ATGGCTTGGCGACGG-
3', how would you have to list the sequence of the complementary
strand, if you want to follow proper convention?

TACCGAACCGCTGCC

CCGTCGCCAAGCCAT

Why can you use the same primers to amplify STRs of different
lengths?

Because all STRs have the same sequence.

Because the flanking regions are identical.

How many STRs does a tetrameric repeat have?

2

3

4

6


Find the TRUE statement(s) about human karyograms.

One can never assign the origin of chromosomes in any karyogram.

One can never assign the origin of somatic chromosomes in any karyogram.

One can always assign the origin of sex chromosomes in a male karyogram.

One can always assign the origin of sex chromosomes in a female karyogram.

One can never assign the origin of sex chromosomes in any karyogram.

Which statement(s) hold true for the TPOX locus?

It has nine different alleles.

The total number of allele combinations is 9 x 9.

It is located on the lower arm of chromosome 2.

It is located on maternal and paternal chromosomes.

It is located in the thyroid proximal gene.


Find the FALSE statement(s) about STRs.

The number of STRs corresponds to the number of alleles.

The number of alleles describes the number of STR variants.

STRs have the same 5' to 3' sequence on both DNA strands.

The frequency of allele combinations varies among different populations.

An individual can have a maximum of two alleles for the same gene locus.

Forensics heavily relies on STR analysis, because:

genetic fingerprints are truly unique.

one can easily amplify STRs by PCR.

the analysis of a single gene locus suffices.

different STR analyses can readily be compared through the patterns they generate.



Homework: chapter 4
Task 1
A human body has approx. 3.72*10^(13) cells. Starting from a single
cell, the fertilized oocyte (or Zygote):
a) how many cell divisions are needed to achieve this number?
b) how many newly synthesized DNA strands does that mean? Show
your calculations.

Character Limit: 1500
Task 2
Polymerase Chain reaction (PCR) allows the amplification of any DNA
template. However, the successful amplification of only the DNA of
interest requires smart designing of the amplification primers. Imagine
you want to amplify short tandem repeats (STRs) as shown in the
figure with the given flanking DNA sequences:

Which of the following primer pairs are you going to use for
amplification of this STR region (remember, DNA sequences are
always given in the 5' to 3' direction)? Briefly explain your answer, by
referring to the PCR principle in general and directionality of
polymerization. Also include one sentence for each of the non-
applicable pairs explaining why they do not yield the desired PCR
product.
Pair 1:
5'-GGCTAGCCTTAGTAACGTTAACTG-3'
5'-AACGTAAGGCCTGTTACGATGACT-3'
Pair 2
5'-GGCTAGCCTTAGTAACGTTAACTG-3'
5'-CCGATCGGAATCATTGCAATTGAC-3'
Pair 3:
5'-GGCTAGCCTTAGTAACGTTAACTG-3'
5'-AGTCATCGTAACAGGCCTTACGTT-3'
Pair 4:
5'-CCGATCGGAATCATTGCAATTGAC-3'
5'-TCAGTAGCATTGTCCGGAATGCAA-3'

Character Limit: 1500
Task 3
STR analyses are also used in paternity testing. In general they same
principles apply as for genetic fingerprinting. In paternity testing,
however, one needs to conclude about the inheritance of genetic
fingerprints.
The figure depicts two STR analyses that were performed in parallel.
Four gene loci were analyzed for the mother, the child and the
presumptive fathers.

Please address the following questions / aspects:
a) Which of the two analyses identifies the genetic father of the child?
b) Name the gene locus based on which you did come to this
conclusion.
c) Briefly explain how the gene locus you picked allows for
identification of the genetic father and exclusion of the second male
tested.

Character Limit: 1500