Assignment 2

1. 5’…….CTTTGCAGAATCTTTATTAGAGTGA 3’
2. DNA primer that will go forward :
5’ ATGGGAAGGGCTCCTTGTTG…3’
DNA primer that will go backward:
5’ TCACTCTAATAAAGATTCTG…3’

3. Total of the volume is not in 25 microliter but it is 27.5 microliter which is over by 2.5
microliter. To know and determine where the error happened, it can be checked by using
titration of the concentration (C1V1=C2V2). Let’s figure it out below:

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Buffer expected final concentration (C2): 1X

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Stocked concentration (C1): 5X

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Final volume added (V2): 25 microliter
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C1V1=C2V2
5X . V1= 1X . 25 microliter
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V1= 25 microliter/ 5 = 5 microliter

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dNTP’s expected final concentration (C2): 200 micromolar

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Stocked concentrartion (C1): 2 milimolar which is equal to 2x10^3

micromolar
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Final volume added (V2): 25 microliter

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C1V1=C2V2
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2x10^3 micromolar.V1=200 micromolar. 25 microliter

Th

V1= (200.25)/2x10^3= 2.5 microliter

DNA primers (both ways) expected final concentration (C2): 0.2 micromolar
sh

Stocked concentration (C1): 5 micromolar

Final volume added (V2): 25 microliter

C1V1=C2V2

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 5 micromolar. V1= 0.2 micromolar. 25 microliter
V1= (0.2x25)/5= 1 microliter

DNA final amount theoretically: 250 nanogram is equal to 0.25 microgram

Stocked amount: 0.5 microgram/microliter
Final amount of required volume: 0.5 microgram/microliter x 100/25 1/microgram = 2
microliter

Taq Polymerase expected total unit: 1.25 unit

Stocked unit: 1 unit/microliter
Final amount of required volume: 1 microliter/unit x 1.25 unit = 1.25 microliter

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Therefore, on that old notebook, there was some errors on the result volume of dNTPs, DNA, and

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also the buffer solution which should be 2.5 microliter, 2 microliter, and 5 microliter. With
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these new result, the required volume that want to be accomplished is satisfied (i.e 25 microliter).
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This is one of the proof: (5+2.5+1+1+2+1.25+11.75) microliter = 25 microliter.
4. The control of this experiment will be having same volume of DNA primer concentration
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which can be as a constant during the proceed, and may result in the expected dependent
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variable. The reversed primer will be 5’ TCACTCTAATAAAGATTCTG…3’ in this

development since it is complementary, the control can not be varied so the result can be
shown as an evidence.
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5. a. y = -0.1598x + 0.9972
b. Lane 1= 0.543 Kb
Lane 2= 0.337 Kb
is

Lane 3= 0.876 Kb
Th

c. The value of 0.337 on Lane 2 is quite strange since it is almost close by 0 and also it is
also considered very small size compare to the other.
sh

d. Lane 3 could contain my PCR reaction since it is not close by 0 size fragment and also
it is one of the largest size that can be found that
6. The formula for this case will be : “20 nucleotides + 6 x amount of the repeated
(“GGGGCC”)” why it can turn out like this since it has stated that “n” will be the

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 amoung of repeated location that can result in Dementia (FTD). Thus, the weight (in bp)
in each individual will be:

Individual A (symptomatic heterozygous 75 and 20 repeated on C9ORF72): (75*6)+20=

470; (20*6)+20= 140
Individual B (asymptomatic homozygous 10 repeated on both C9ORF72) : (20*6)+20=
140; (20*6)+20= 140
Individual C: (symptomatic homozygous 130 repeated on both C9ORF72):
(130*6)+20= 800; (130*6)+20= 800

It can be resulted in PCR in Lane:

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Individual A (2 spots): 140 bp and 470 bp

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Individual B (1 spot): 140 bp

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Individual C (1 spot): 800 bp
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