Page 1 of 11
PRACTICE THEORETICAL BIO3151 EXAM
Part I : BIOINFORMATICS
1. From which organism does the following sequence come from?
CGAAGACCACGATTAGCTCTTCCCATGGCTTCTTCACTGAAGAAGCTTATCTCA
AGCTTTCTACTTGTCTTATACTCCACCACCATCATTGTTGCTTCATCAGAATCTC
GATGTAGGCGTTTTAAATCGATCATCAGCTTTGGTGATTCCATCGCAGACACCG
GAAATTATCTCCATCTCTCTGATGTCAATCACCTTCCTCAATCCGCCTTTCTTCC
TTATGGCGAAAGCTTCTTCCATCCTCCCTCCGGTCGTGCCTCTAATGGCCGTCT
CATCATCGACTTCATTGGTAGCTATTTTTTCATCTCTATAGGCAAAACCACCTGT
TATCATTTGCTCTAAGTCTTTCTTACGTGAACAGCAAGTTATACTTCTTCCTGATC
TTTTTTTCTTATGTTGTCTTACTATTTGAACAGCCGAATTCTTGGGACTACCATAC
GTACCGCCTTACTTTGGATCCCAAAACGTTAGCTTCGAACAAGGGATC
2. Give the accession number of a protein orthologue of this sequence from turnip
(Brassica napus).
Use the following sequences to answer the questions that follow:
CGAAGACCACGATTAGCTCTTCCCATGGCTTCTTCACTGAAGAAGCTTATCTCAAGC
TTTCTACTTGTCTTATACTCCACCACCATCATTGTTGCTTCATCAGAATCTCGATGTA
GGCGTTTTAAATCGATCATCAGCTTTGGTGATTCCATCGCAGACACCGGAAATTATC
TCCATCTCTCTGATGTCAATCACCTTCCTCAATCCGCCTTTCTTCCTTATGGCGAAA
GCTTCTTCCATCCTCCCTCCGGTCGTGCCTCTAATGGCCGTCTCATCATCGACTTC
ATTGGTAGCTATTTTTTCATCTCTATAGGCAAAACCACCTGTTATCATTTGCTCTAAG
TCTTTCTTACGTGAACAGCAAGTTATACTTCTTCCTGATCTTTTTTTCTTATGTTGTCT
TACTATTTGAACAGCCGAATTCTTGGGACTACCATACGTACCGCCTTACTTTGGATC
CCAAAACGTTAGCTTCGAACAAGGGATC
P1: CGGAACATACATAAGCCAAAATAG
P2: GAAGTGCACTTACCGTTGCTGTG
P3: CAGCTTGATGGCATTGAACAG
P4: TCCTTGCCGCGCATGTCCGAC
a. Which primer or primers could be used for a gene specific reverse transcriptase
reaction of the longest protein coding mRNA encoded by this sequence?
b. What is the identity of the translated protein from the longest protein coding mRNA
encoded by this sequence?
Page 2 of 11
PRACTICE THEORETICAL BIO3151 EXAM
3. Use the sequence NM_167140 to answer the following questions:
a. Which of the following primers could be used to amplify at least 200bp of this
sequence?
P1: CGGAACATACATAAGCCAAAATAG
P2: GAAGTGCACTTACCGTTGCTGTG
P3: CAGCTTGATGGCATTGAACAG
P4: TCCTTGCCGCGCATGTCCGAC
b. Which of the restriction sites in list 1 could be added to the primers for the directional
cloning of the PCR product within the restriction sites of the multiple cloning site
indicated in list 2:
List 1:
List 2:
Bcl I : T▼GATCA
Bmt I : GCTAG▼C
ClaI : AT▼CGAT
Rsa I : GT▼AC
Sal I : G▼TCGAC
Bam HI : G▼GATCC
Pst I : CTGCA▼G
Sma I : CCC▼GGG
Xba I : T▼CTAGA
Xho I : C▼TCGAG
Page 3 of 11
PRACTICE THEORETICAL BIO3151 EXAM
Part II : RESTRICTION MAPPING
A recombinant plasmid was generated as follows: PCR was used to amplify a region of
the yeast genome with a pair of “Forward” and “Reverse” primers which had the
restriction sites Pst I and Pvu II added to their 5’ ends. Following digestion of the
amplicon with Pst I and Pvu II, it was ligated into the pUC vector digested with Pst I and
Sma I.
Pst I
Pvu II
Sma I
To verify the identity of the recombinant plasmid, restriction digests followed by agarose
gel electrophoresis were performed. The gel and the corresponding standard curve of
the molecular size markers are illustrated. The values next to each band indicate the
migration distances in mm.
U
E
H
X
X+E
100 000
2.0
3.0
2.1
3.0
2.1
4.3
5.4
3.7
6.8
U: Uncut
E: EcoRI
H: HindIII
X: XhoI
10
Size (bp)
10 000
1 000
0
5
10
15
Migration distance (mm)
20
25
Page 4 of 11
PRACTICE THEORETICAL BIO3151 EXAM
1. Which lane or lanes demonstrate partially undigested plasmid DNA?
2. Which lane or lanes illustrate partially digested DNA?
3. Which lane or lanes illustrate complete digests?
4. What is the most accurate approximate size of the insert?
5. The PCR product inserted in the pUC vector was used as a probe for a Southern
analysis. Which bands in lane H would hybridize to the probe?
6. How many times does “H” cut within the insert?
7. What fragment size(s) would be predicted from a digest of the recombinant plasmid
with Pst I and SmaI?
8. What is the probable distance from the Eco RI going towards the insert to the Xho I
restriction site?
9. Given that all the digests were done with 0.1µg of DNA, how much DNA is probably
represented by the band with a migration distance of 6.8 mm in lane “X + E”?
10. Which band in lane “U probably represents supercoiled plasmid DNA?
Page 5 of 11
PRACTICE THEORETICAL BIO3151 EXAM
PART III : CALCULATIONS
1. Solution "A" has 1.20 µg of DNA per µl. Solution "B" has 3.10 µg of DNA per µL. If
you combine 34 µl of solution "A" with 19 µl of solution "B", what is the DNA
concentration in µg/µl of the new solution? Round the answer to 2 decimal places.
2. The concentration of a salt solution is 3.8 mg/ml. 124 ml of this solution is placed in
a beaker and allowed to evaporate until 92 ml remains. What is the new salt
concentration? (Assume that water evaporates but that the salt does not.) Round
the answer to 1 decimal place.
3. You have 3g of NaCl [MW=58]. You need a 1M solution. How much solution can
you make?
4. You have a DNA solution with an absorbance at 260nm of 0.5. How much of this
DNA solution and water would you need to prepare 1 ml of 10 μg/ml solution?
5. You need to administer 10mg of epinephrine to a patient to save her life. How much
of a 200 μg/ml solution of epinephrine would you administer?
6. You have a solution of 1M CrCl3 and 0.5M K2SO4. How many moles of Cl- are there
for each mole of K+?
7. A PCR reaction is setup with an equal quantity in µg of a double stranded DNA
template of 10 000 base pairs and two single stranded primers (P1 and P2) which
are 20 and 25 bases long respectively. What is the molar ratio between each of the
primers and their respective templates? (Ex. P1:P2:template = 1:1:1)
8. A solution is prepared by mixing 24 parts of 1M glucose with 5 parts of 4.8M NaCl
and 1 part of water. What are the concentrations of glucose and NaCl in the final
solution?
9. Vinegar is sold as a 5% (v/v) solution of acetic acid in water. What volume of water
should be added to 15 mL of pure acetic acid (100%) in order to generate a 5% (v/v)
solution of acetic acid?
10. What is the molarity of a 60% (m/m) solution of ethanol (MW: 46.0) in water of which
the density is 0.8937 g/mL?
11. What are the molar concentrations of iron (Fe) and of sulfate (SO4) in a solution of
0.450M Fe2(SO4)3?
12. A car’s antifreeze mixture is made by mixing equal volumes of ethylene glycol (d =
1.114 g/mL, molar mass 62.07 g/mol) and water (d = 1.000 g/mL) at 20.0 °C. The
density of the solution is 1.070 g/mL. Express the concentration of ethylene glycol
as:
(a) volume percent
(b) mass percent
(c) molarity
Page 6 of 11
PRACTICE THEORETICAL BIO3151 EXAM
13. A solution is prepared by dissolving 28.0 g of glucose in 350 g of water. The density
of the resulting solution was 1.54. What is the molarity of the solution? (MW of
glucose: 180g/mole)
14. How many mL of a 0.375 M solution can be made from 35.2 g of Ca3(PO4)2
(MW=310 g/mole)?
15. 315 mL of a 0.525 M solution of NaCl (MW=58.4 g/mole) is mixed with 188 mL of a
0.295 M solution of NaCl, then an additional 166 mL of water is added. What is the
final molarity of solution?
16. The term proof is defined as twice the percent by volume of pure ethanol in solution.
Thus, a solution that is 95% (by volume) ethanol is 190 proof. What is the molarity of
ethanol in a 92 proof ethanol/water solution?
Density of ethanol 0.80 g/ml
Density of water 1.0 g/ml
Molecular wt. of ethanol 46
17. 5.00 g of potassium chloride (KCl; MW: 74.5) is dissolved in enough water to
make 250.0 mL of a solution with a density of 1.14g/mL. What is the
concentration of the solution in % (m/m), % (m/V), and molarity?
18. 650 mL of a 2M solution of compound X is totally evaporated leaving 120g of
compound X. What is the molecular weight of compound X?
Page 7 of 11
PRACTICE THEORETICAL BIO3151 EXAM
PART IV PROBLEMS
1. Below are the DNA fingerprints of 5 people including: a child (C), the mother (M) ,
and three potential fathers of the child (F1, F2, F3). In this experiment, three different
loci (A, B, and C) containing a VNTR were amplified from the genomes of each
individual.
VNTR A
VNTR B
VNTR C
a. Who can be eliminated as the biological father of child "C"?
b. Which man or men can be the biological father of child “C”?
c. A 5Kbp PCR amplified fragment includes two RFLPs. The restriction sites of the
RFLPs are labelled “X” and “Y”. The mutant alleles of "X" and “Y” cannot be cut, but
the normal alleles can. The positions of the restriction sites on the PCR fragment
are as follows:
X
Y
2Kbp
2Kbp
1Kbp
You do a digest of the PCR products from two individuals. Based on the gel below
and the map above, what are the phenotypes of each individual for each allele:
homozygous mutant (-/-), heterozygous (+/-), or homozygous normal (+/+)? Indicate
the correct combination from the table below. (Ex. AE)
Individual 1
Individual 2
6Kbp
5Kbp
4Kbp
3Kbp
2Kbp
1Kbp
A
B
C
D
Individual 1:
X(-/-) Y(-/-)
X(+/-) Y(+/+)
X(-/-) Y(+/-)
X(+/+) Y(-/-)
Individual 2:
E X(-/-) Y(-/-)
F X(+/-) Y(+/+)
G X(-/-) Y(+/-)
H X(+/+) Y(-/-)
d. In the case of individual 1, how would the results observed on the gel be different if
the genomic DNA had been digested before performing the PCR?
Page 8 of 11
PRACTICE THEORETICAL BIO3151 EXAM
2. Southern hybridizations were performed on Bam HI digested genomic DNA
from either humans or chimpanzees. The hybridizations were performed with
increasing concentrations of formamide (lanes 1-6) and two DNA probes
derived from a given gene.
1 2 3 4 5 6
1 2 3 4 5 6
Probe 1
1
2
3
4
5
6
1
2
3
4
5
6
Probe 2
Chimpanzee genomic
2a. Probes 1 and 2 represent DNA from the same DNA
organism. From which
Human genomic DNA
organism were the probes most probably derived? Justify your answer.
2b. BamHI cuts how many times within the region of the human genome spanned
by probe 1?
2c. BamHI cuts how many times within the region of the chimpanzee genome
spanned by probe 1?
Page 9 of 11
PRACTICE THEORETICAL BIO3151 EXAM
3.
3’ TGCTAGCTGAAATCGATCCG………………………………CCGGCCCCGCTGGATCCAG 5’
5’ ACGATCGACTTTAGCTAGGC………………………………GGCCGGGGCGACCTAGGTC 3’
A probe was synthesized from the above sequence (“My Gene”) by performing a PCR
reaction with the following single primer: 5’ ACGATCGACTTTAGCTAGG 3’, in the
presence of dig conjugated dUTP. Below is a diagram showing a northern blot
hybridization with this probe to RNA isolated from the thymus. The membrane was also
probed at the same time with a second probe against actin; a house keeping gene. The
time indicates how long the cells were incubated in the presence of a drug before the
RNA was isolated.
My Gene
Actin
3a. According to the results obtained, which of the above indicated strands of “My Gene”
was read (transcribed) by the DNA dependent RNA polymerase? (Top or Bottom?)
3b. What is the effect of the drug on the expression of “My Gene”? Justify your answer.
.
3c. You wish to repeat this experiment by performing a gene specific RT-PCR. Indicate
the sequence of a 10 base primer that could be used for the reverse transcription of
“My Gene”. Make sure to label your ends.
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PRACTICE THEORETICAL BIO3151 EXAM
4.
5’TGCTAGCTGAAATCGATCCG…………………………………………CCGGCCCCGCTGGATCCAG3’
3’ACGATCGACTTTAGCTAGGC…………………………………………GGCCGGGGCGACCTAGGTC5’
A probe was synthesized from the above sequence by performing a PCR reaction with
the following primer: TAGCTGAAATCGATCC in the presence of radioactive nucleotides.
A Southern (panel “A”) and a Northern (panel “B”) were then hybridized with the probe
generated.
A.
F
R
A
B.
1
2
← RNA of interest
←House keeping RNA
Panel A: Southern of genomic DNA digested with “F”, “R”, or “E”.
Panel B: Northern of total RNA isolated from cells in conditions “1” and “2”.
a. If the sequence represented by the probe is only found once within the genome, how
many times do each of the enzymes “F”, “R” and “A” cut within the probe’s
sequence?
b. According to the results of the Northern, the sequence of which of the two strands
must have been read to generate the mRNA? (Top or Bottom)
c. According to the Northern, does either of the conditions result in the induction of
transcription? If so, which one? Briefly justify.
d. Indicate in the table below which of the single stranded sequences indicated in the
column “probe” could hybridize to the target sequence at the indicated temperatures.
Use the symbols “+” and “-“ to indicate whether or not hybridization would occur.
Probe
ATCATTTGTCCGATGCTGGATC
ATCACCCGTCCGATGCTGGATC
ATCACTTGTAAAATGCTGGATC
Target sequence: GATCCAGCATCGGACAAATGAT
Hybridization Temperatures
45
56
65
Page 11 of 11
PRACTICE THEORETICAL BIO3151 EXAM
PART V: THEORY
For each group of statements, indicate in the space provided which one (s), are true. If
none are true indicate "0":
I.
Both plasmid isolation methods as well as the RNA isolation method used in this
lab make use of a strong alkali to lyse the cells.
II.
The Qiagen plasmid isolation method uses chromatography to remove proteins
whereas the alkaline lysis procedure uses differential sedimentation.
III.
Both plasmid isolation methods as well as the RNA isolation method used in this
lab make use of differential sedimentation to obtain the desired nucleic acids.
I.
Isoschizomers are different enzymes from a same bacterial species which
recognize and cut the same sequence.
II.
Neoschizomers necessarily generate compatible ends.
III.
Ends generated by two isocaudaumers can be ligated to one another.
a.
b.
I.
Given that ethidium bromide binds DNA by electrostatic interactions one would
expect the ethidium bromide to migrate towards the negative electrode.
II.
Supercoiled plasmid DNA migrates through agarose with the least resistance.
III.
Denatured RNA would be expected to migrate faster through agarose than nondenatured RNA.
I.
A selectable marker is included in cloning vectors to increase the number of cells
which uptake a plasmid.
II.
All vectors are circular DNA molecules.
III.
All vectors are plasmids and all plasmids are vectors.
c.
d.
I.
The Tm of nucleic acids decreases with increasing concentrations of positive
ions.
II.
Formamide and formaldehyde decrease the Tm of nucleic acids.
III.
At high stringency it is more likely to obtain hybridization of a sequence with a
high %GC rather than a high %AT.
e.