Module 3 Section 1-3

Question 1

Purines

Answer 1

Adenine, guanine

Question 2

Pyrimidines

Answer 2

Cytosine, Thymine, Uracil

Question 3

Chargoff’s rules

Answer 3

1. base composition of DNA varies from species to species
2. DNA samples from different tissues of the same species have the same base composition
3. DNA base composition does not change with age, nutritional state, environment
4. (A=T, G=C)

Question 4

H-bonding in DNA relative strengths

Answer 4

G-C bond stronger b/c 3 H-bonds (A-T has 2)

Question 5

DNA Major grove

Answer 5

-Where nucleotide sequence is primarily read by DNA binding proteins

Question 6

DNA length per turn

Answer 6

10.5 base pairs in a complete helix turn

Question 7

When studying a virus, discovered that it is 19% adenosine, 32% thymidine. What does this tell you about the virus?

Answer 7

• is DNA, has thymidine
• adenosine is not equal to thymidine, so it cannon be double stranded DNA
• therefore is ssDNA

Question 8

Functions of DNA

Answer 8

• long term storage for genetic info
• template for DNA replication
• coding for proteins and functional RNAs

Question 9

DNA vs RNA stability

Answer 9

• DNA very stable, RNA/proteins are not (have to be environmentally sensitive)
• RNA unstable by design, hydroxide anions deprotonate 2’OH, which attacks phosphodiester bonds
• 2’ and 3’ cyclic monophosphates converted to 2’ and 3’ monophosphates

Question 10

RNA 2˚ structure

Answer 10

• single stand, folded back on self
• hydrophobic bases stacked within helix
• fully/partially base paired
• antiparallel
• helices, internal loops/bulge (dsRNA separates due to lack of watson-crick base pairing), hairpin loop (most common, unpaired bases at loop)
• actions of functional RNA governed by the 2˚ structure
• can have non-watson-crick pairings (ie. G-T)
• can have base-triple interations (3-BP interaction)

Question 11

What chemical interaction is NOT involved in stabilizing the transition of ssDNA into a double helical structure?

Answer 11

• Phosphodiester bond

- involved in the structure of DNA (backbone), but the helical structure is NOT stabilized by covalent interactions

Question 12

Interactions that result in DNA double helix stabilization

Answer 12

H-Bonding
-A-T has 2 H-bonds, G-C has 3
-areas where helicase has to act usually has A-T rich regions due to weaker interaction between A and T
Hydrophobic+Van der Waals
-bases are hydrophobic, align to inside of mlc
Ionic interactions
-backbone highly neg. charged at neutral pH, electrostatic repulsion
-addition of salt neutralizes -ve charge

Question 13

What would be the consequence of adding salt to a solution of dsDNA?

Answer 13

It would take a higher temp to melt. The helix will be stabilized by the cations

Question 14

To promote DNA melting (denaturation)

Answer 14

• increase temp
• reduce salt
• add organic solvents (ie. urea)
• add DNA binding proteins (ie. helicase)

Question 15

Quantification of nucleic acids

Answer 15

• nitrogenous bases absorb light (conjugated)
• use beer’s law (A=ecl)
• order of UV absorption: free dNTPs>ssDNA>dsDNA

Question 16

Melting point (Tm)

Answer 16

-temp where half the DNA is denatured

Question 17

Internal factors the influence Tm

Answer 17

-Base composition (G-C stronger than A-T)
Tm=0.41x(%G+C)+69.3˚C
-true for long DNA segments
-Salt concentration of 0.2M

Question 18

External Factors that influence Tm

Answer 18

• High pH (causes deprotonation of bases, reduced H-bonding)
• addition of organic solvents decreases Tm
• addition of salt increases Tm

Question 19

DNA hybridization

Answer 19

• separated DNA strands can renature with other polynucleotides sharing complementary sequences
• ie. cross species, DNA-RNA, DNA-DNA
• the stability of the hybrid depends on the stringency of the conditions

Question 20

Stringency

Answer 20

• The strictness with which Watson-Crick pairing is required for nucleic acid hybridization under specific conditions of temp, pH, [Salt], etc.
• high stringency: hybridization occurs only when the two strands are highly compatible