Exp 4.1: Nucleic Acids (NA)

Question 1

Identify:
repository of genetic information

Answer 1

nucleic acids

Question 2

Enumerate:
2 basic structural forms of nucleic acids

Answer 2

1. deoxyribonucleic acid (DNA)
2. ribonucleic acid (RNA)

Question 3

Identify which structural form of NA:
genetic material

Answer 3

DNA

Question 4

Identify which structural form of NA:
provides template for RNA transcription

Answer 4

DNA

Question 5

Identify which structural form of NA:
carriers of genetic information for protein translation

Answer 5

RNA

Question 6

Identify:
biopolymers of nucleotide

Answer 6

nucleic acids;
(nucleotides form nucleic acids)

Question 7

Enumerate:
basic components of nucleic acids

Answer 7

1. pentose sugar
2. heterocyclic nitrogen bases
3. phosphate

Question 8

Identify which basic component of NA:
attached by a N-C glycosidic to the 1’-carbon atom of the sugar component

Answer 8

heterocyclic nitrogen base

Question 9

Identify:
nucleotide’s number of phosphate groups

Answer 9

1, 2, or 3

Question 10

Identify:
reason for acidic and anionic character

Answer 10

phosphate groups

Question 11

Identify:
where phosphate is attached

Answer 11

attached to the 5’-carbon atom of the sugar component

Question 12

Identify:
form the symmetrical backbone of the nucleic acid with the 5’ end of one sugar always linked through a phosphate molecule to the 3’ end of the adjacent sugar

Answer 12

sugar-phosphate linkages

Question 13

Enumerate:
interactions responsible for NA rigid molecular configuration

Answer 13

1. phosphodiester bonds
2. hydrogen bonds
3. Van der Waal’s forces

Question 14

Identify the interaction responsible for NA rigid molecular configuration:
links nucleotides in the polynucleotide chain

Answer 14

phosphodiester bonds

Question 15

Identify the interaction responsible for NA rigid molecular configuration:
involved in the complementary base pairing in DNA (& RNA i.e. DNA: RNA hybrids)

Answer 15

hydrogen bonds

Question 16

Identify the interaction responsible for NA rigid molecular configuration:
base stacking - stacking interactions

Answer 16

Van der Waal’s forces (pi-pi complexation)

Question 17

Identify:
a process of DNA purification from a sample using a combination of physical and chemical methods

Answer 17

DNA isolation

Question 18

Enumerate:
methods used for DNA isolation are dependent on?

Answer 18

1. source of sample
2. age of sample
3. size of sample

Question 19

Identify
DNA isolation’s aim

Answer 19

to separate the DNA present in the nucleus of the cell from other cellular components

Question 20

Identify:
major component of all cells;
DNA source

Answer 20

nucleic acid
(5-15% of cell’s dry weight)

Question 21

Identify:
choice of DNA sample

Answer 21

tissue containing cells with high nuclear-cytoplasmic volume ratio

Question 22

T/F:
DNA concentration is high in the nucleus

Answer 22

true

Question 23

Enumerate:
common DNA sources

Answer 23

1. whole blood
2. hair
3. sperm
4. bones
5. nails
6. tissues
7. blood stains
8. saliva
9. buccal swabs
10. urine

Question 24

Enumerate:
organs rich in nucleic acids

Answer 24

1. spleen
2. liver
3. thymus
4. pancreas

Question 25

T/F: brain is the best source of DNA

Answer 25

false; thymus

Question 26

T/F: brain and muscle tissues have lower DNA concentration

Answer 26

true

Question 27

T/F: other DNA sources can be archived tissue samples like frozen blood or tissue exhumed from bones, FFPE

Answer 27

true

Question 28

Enumerate: DNA isolation applications

Answer 28

1. paternity testing 2. forensics

Question 29

T/F: It is necessary to isolate DNA because presence of proteins, lipids, polysaccharides and other organic/ inorganic components in the DNA preparation can interfere with DNA analysis method.

Answer 29

true

Question 30

T/F: It is necessary to isolate DNA because presence of other components also reduce the quality of DNA leading to its shorter storage life.

Answer 30

true

Question 31

Enumerate: factors to consider in nucleic acid isolation

Answer 31

1. high content of DNA and RNA 2. low DNAse and RNAse activity 3. nucleic acids isolated from source are readily obtainable in pure form 4. large size and fragile nature of DNA 5. RNA is less stable (compared to DNA)

Question 32

Enumerate reasons: why the large size and fragile nature of DNA is considered in NA isolation

Answer 32

1. difficulty in isolating intact and undamaged DNA 2. high molecular weight and intact DNA vs. sheared or fragmented DNA

Question 33

Enumerate reasons: why the stability of NA is considered in NA isolation

Answer 33

RNA: less stable 1. single-stranded molecule 2. presence of 2--hydroxyl group

Question 34

Enumerate: experimental (physical & chemical) factors

Answer 34

1. pH: extremes in pH 2. temperature: heating 3. ionic strength 4. cellular conditions 5. mechanical stress 6. storage of DNA/ RNA

Question 35

Identify the pH range: hydrogen bonding between complementary strands in stable between?

Answer 35

pH 4-10

Question 36

Identify the pH range: DNA is hydrolyzed at what pH

Answer 36

pH less than 3 or greater than 12

Question 37

Identify the temperature range: most DNA begins to unwind in the range?

Answer 37

80°-90°C

Question 38

Identify the temperature: phosphodiester bonds and β-N, C-glycosidic bonds are stable up to?

Answer 38

100°C

Question 39

Identify: DNA ionic strength and solubility

Answer 39

DNA is most stable in ↑[salt]; insoluble ↓[salt solutions]

Question 40

Explain: reason behind DNA stability and solubility in salt concentrations

Answer 40

salt concentrations of less than 0.1M weaken hydrogen bonding between complementary strands

Question 41

Enumerate mechanisms: cellular conditions as an experimental factor

Answer 41

1. lysis or disruption of the biological membrane release DNA when the tissue is homogenized 2. presence of nucleases

Question 42

Identify: grinding, shaking, stirring, squirting the solution through narrow orifices and other disruptive procedures may cause cleavage (shearing or scission) of the DNA chains

Answer 42

mechanical stress

Question 43

T/F: mechanical stress damages DNA secondary structure but increases DNA length

Answer 43

false; does not damage reduces DNA length

Question 44

T/F: Purified DNA is best kept in solution.

Answer 44

true

Question 45

Explain: Why is purified DNA best kept in solution?

Answer 45

If precipitated DNA is kept dry, it will undergo denaturation.

Question 46

Enumerate: general steps in DNA isolation

Answer 46

1. homogenization 2. dissociation and denaturation of the nucleoprotein (deproteinization) 3. precipitation 4. purification of nucleic acid

Question 47

Identify the process: lysis or disruption of biological membranes

Answer 47

homogenization

Question 48

Enumerate: 2 ways of hoogenization

Answer 48

1. mechanical: mincing, grinding, sonication 2. chemical: detergents & chaotropic agents