Conformations of DNA and DNA denaturation

Question 1

B Form of DNA

Answer 1

Conformation that the sodium salt of DNA assumes under conditions of relative high humidity ~92% considered “common form”

Base pairs perpendicular to the helical axis.
10 bp per helical turn
pitch of 34 angstroms (pitch=one turn of helix)

Question 2

A Form of DNA

Answer 2

Humidity reduced to ~75%

Base airs tilted 20 degree angle
11 bp per helical turn
pitch of 28 angstroms

*DNA RNA hybrids

Question 3

Z DNA

Answer 3

Composed of alternating purines and pyrimidines =left handed helix
backbone appeared to zigzag
Tilt of 9 degrees
12 bp per helical turn
pitch of 46 angstroms

Question 4

DNA denaturation

Answer 4

melting
presence of organic solvents DMSO
high pH

Question 5

How does the lowering of the salt concentrations in solution aide DNA denaturation?

Answer 5

Ions are removed that shield the negative charges on DNA strands from each other. The repulsive forces of the negative charges are such that DNA can denature at low temperatures.

Question 6

DNA Melting

Answer 6

DNA denaturation occurs when the two strands come apart.

Question 7

Melting Temperature (Tm)

Answer 7

Temperature at which the DNA strands are HALF denatured

Question 8

Hyperchromic Shift

Answer 8

When two strands separate the quenching of the absorbance is reduced and the absorbance at 260nm INCREASES 30-40%

Question 9

Measurement of DNA strand separation

Answer 9

Absorbance of DNA solution at 260nm.

Increasing the distance between the bases increases this absorbance.

Question 10

How does the GC content of DNA effect its stability, Tm, and density?

Answer 10

Increase GC content = linear Increase Tm and density.

GC have a 3rd H-bond that effect DNA Tm
Natural occurring DNA GC content of 22-73%

Question 11

Renaturation or Annealing

Answer 11

reuniting of DNA strands

Question 12

What are the 3 factors that contribute to the renaturation efficiency?

Answer 12

Temperature
DNA Concentration
Time

Question 13

How does temperature effect renaturation?

Answer 13

A Tm of =Tm-25C is best for renaturation to occur.

It is low enough that it does not promote melting but high enough to allow rapid diffusional movement of DNA and weaken transient bond between sequence mismatches.

Question 14

How does rapid cooling effect DNA renaturation?

Answer 14

Rapid cooling of denatured DNA prevents renaturation and is called quenching.

Question 15

How does DNA concentration effect DNA renaturation?

Answer 15

The higher the DNA concentration the faster the renaturation.

Question 16

How does the amount of time effect DNA denaturation?

Answer 16

The longer the time allowed for annealing the more will occur.

Question 17

Fourth Factor that contributes to DNA renaturation?

Answer 17

DNA Complexity.

The more complex the DNA the slower and less efficient renaturation.

Question 18

What is the C0t term combine and what are the components measured in?

Answer 18

C0t combines DNA concentration and time in DNA renaturation studies. Concentration multiplied by time.

DNA concentration C0 expressed in moles of nucleotides per liter.

Time t expressed in seconds.

Question 19

What is “Half C0t”?

Answer 19

C0t is usually expressed as C0t1/2 or half C0t and it represents the C0t at which a DNA sample is half annealed.

Question 20

Hybridization

Answer 20

A process in which a DNA strand anneals with a complementary strand of RNA to form a HYBRID duplex

Question 21

DNA size range?

Answer 21

From a few kilobases to hundreds of megabases.

Question 22

What is TEM?

Answer 22

Transmission electron microscopy.
It is a convenient means to estimate the size of small DNAs and also shows if DNA is linear, circular, or supercoiled.

DNA is more commonly estimated by electrophoresis.

Question 23

The C-Value Paradox

Answer 23

One would guess that the more DNA an organism has the more complex the organism and the more genes it has. there is a rough correlation between DNA content and number of genes BUT in a number of cases the correlation breaks down.

For Example: mouse and human haploid cells contain 100X more DNA than yeast haploid cells but frogs have 7X and Lilys 100X more than humans.

The C-Value paradox explained by extra DNA in some organisms.

Question 24

Paradox

Answer 24

A paradox is a statement that apparently contradicts itself and yet might be true.

Question 25

C-value

Answer 25

DNA content per haploid cell

Question 26

what is a reliable method to estimate the number of genes in an organism?

Answer 26

to estimate the number of different mRNAs.