EXAM 3, BICH 410 (MWF 1:50 PM), Monday, Nov. 13, 1995


Write your name on each page. Write concise answers to demonstrate effectively your mastery of the subject material. Show your work in order to receive partial credit where applicable.

1) (10 pts) (A) What is the difference between a genomic library and a cDNA library?
(B) Which type of library contains more different kinds of sequences? Why?
(C) What special enzyme must be used to prepare a cDNA library?

2) (10 pts) Draw the structure of 2-prime,3-prime-ddGTP.

3) (10 pts) Given the following sequence of single-stranded RNA. Answer the questions below.

5prime-GGAUAAUCAUUUGCAAUUGCAGGCUGGCCCUGCUUUAACAAGUUAA-3prime

(A) Write the sequence of a 15 nucleotide DNA that would form the most stable hybrid with this RNA sequence. Make sure you denote the polarity of this strand.

(B) Using the 15 nt. oligonucleotide from part (A) as a primer for DNA synthesis from the complementary RNA strand, write the sequence of DNA produced from the primer when only the nucleotides ddGTP, dTTP, dATP and dCTP are added to the reaction. Show polarity.

(C) What potential problem might exist when using the primer in (A) as a probe for hybridization to this RNA sequence (Hint: Carefully consider the sequence of the 15 nt oligonucleotide).

4) (15 pts) (A) Draw the structure of the following hypothetical disaccharide:
O-D-3-deoxyribofuranosyl-ß(1-6)-D-glucopyranoside.

(B) Without re-drawing the entire structure, indicate what compound is produced from this disaccharide after treatment with Fehlings reagent (Cu2+). (that is, demonstrate what part of the molecule is transformed and to what functional group(s))
(C) Write the name of a disaccharide using the same two monosaccharides as in (A) containing a glycosidic linkage that would result in a nonreducing sugar.

5) (13 pts) A relaxed, circular, double-stranded DNA molecule in B-form contains 6300 bp.
(A) What is the linking number (Lk) of this relaxed DNA molecule?
(B) If DNA gyrase (plus ATP) introduced 12 negative supercoils, what would be the values for Lk, W and T?
(C) What is the superhelical density for the DNA molecule in (B)?
(D) If the starting, relaxed DNA molecule was in Z-form, what would be the linking number?

6) (12 pts) Use the table of restriction enzyme recognition sequences (see last page of this exam) to answer the following questions.
(A) Write the sequence of both strands at the end of a fragment of DNA cut by PstI. Indicate the polarity (5prime, 3prime) of each strand. What is this type of end called?
(B) Restriction fragments from which two different enzymes listed in the table are capable of being joined by DNA ligase? Write the sequence of both strands of the junction after ligation of these fragments. Indicate the polarity of each strand.
(C) The double-stranded oligonucleotide whose sequence is given below is ligated to the end of another fragment prepared by cutting with EcoRI. After ligation, can EcoRI be used again to cut out the oligonucleotide? With a diagram explain your answer.

5prime---------CTTGGCGCAC--------3prime
3prime-AATTGAACCGCGTGTTAA-5prime

7) (30 pts) Answer the following questions.
(A) Without using an enzyme, what simple condition can be used to selectively degrade RNA, but not DNA?
____________________________________________________________
What modification to the carbohydrate portion of RNA (not 2prime-deoxy) would prevent this degradation from occurring?
________________________________________________
(B) Which blood group carbohydrate structure is the smallest and not antigenic? _____________.
An individual homozygous for this blood group antigen can accept blood from what other types?
_____________________________
(C) What is the chemical basis for the difference between cellulose and amylose?
____________________________________________________
(D) Write the long, formal name (describing the stereochemistry and glycosidic linkage) for sucrose.
____________________________________________________
(E) The carbohydrate-containing structure that is contained in a bacterial cell wall is called
_____________________. An enzyme that degrades this structure is ________________.
(F) Deamination of 5-methylcytosine yields __________________________.
(G) A nucleosome core particle contains ___________ bp of DNA.
The histone composition of the core particle is:
________________________________________________________________.
(H) A peak of absorption of UV light by nucleic acids occurs at what wavelength? ____________.

Which absorbs the most UV light? (Circle correct answer)
i) double-stranded DNA
ii) single-stranded DNA (both strands present)
iii) single-stranded DNA (only one strand from the double-stranded molecule remains, but the volume of the sample is unchanged)


Restriction Enzyme Recognition Sequences (for Prob. 6)

TaqI
5prime-TCGA-3prime
3prime-AGCT-5prime

PstI
5prime-CTGCAG-3prime
3prime-GACGTC-5prime

XhoI
5prime-CTCGAG-3prime
3prime-GAGCTC-5prime

EcoRI
5prime-GAATTC-3prime
3prime-CTTAAG-5prime

SalI
5prime-GTCGAC-3prime
3prime-CAGCTG-5prime

HindIII
5prime-AAGCTT-3prime
3prime-TTCGAA-5prime