Ch 6 DNA and Biotechnology

Question 1

DNA and RNA

Answer 1

Polymers that together create molecules integral to life

DNA is recorded from 5’-3’

Question 2

Nucleosides

Answer 2

Five-carbon sugar (pentose) bonded to a nitrogenous base formed by covalently linking the base to C-1’ of the sugar
Comprise nucleotides

Question 3

Nucleotides

Answer 3

Formed when one or more phosphate groups are attached to C-5’ of a nucleoside
Often named according to number of phosphates present.
High energy because of energy of repulsion between negative charges on phosphate groups
Building blocks of DNA

Question 4

Ribose

Answer 4

Five-carbon sugar (pentose) in a ring with O between C-1 and C-4, another C and OH off of C-4 and Ohs

Question 5

Deoxyribose

Answer 5

“Deoxygenates” C-2 on pentose to just H.

Question 6

Base Adenine

Answer 6

Nucleoside: Adenosine (deoxyadenosine)
Nucleotides: AMP (dAMP), ADP (dADP), and ATP (dATP)

Question 7

Guanine Base

Answer 7

Nucleoside: Guanosine (deoxyguanosine)
Nucleotides: GMP (dGMP), GDP (dGDP), GTP (dGTP)

Question 8

Cystosine

Answer 8

Nucleoside: Cytidine (deoxycytidine)
Nucleotides: CMP (dCMP), CDP (dCDP), CTP (dCTP)

Question 9

Uracil

Answer 9

Nucleoside: Uridine (deoxyuridine), UMP (dUMP), UDP (dUDP), UTP (dUTP)

Question 10

Thymine

Answer 10

Nucleoside: (deoxythymidine) no thymidine because it appears almost exclusively in DNA
Nucleotides: (dTMP), (dTDP), (dTTP)

Question 11

Purines

Answer 11

Aromatic, Contain two rings in their structure

The two found in nucleic acids are adenine (A) and guanine (G) both found in both RNA and DNA

Question 12

Pyrimidines

Answer 12

Aromatic, Contain only one ring in their structure

The three are cytosine (C), Thymine (T), and uracil (U)

Question 13

Aromatic

Answer 13

Unusually stable ring system that adheres to these:

1. Compound is cyclic
2. Compound is planar
3. Compound is conjugated (has alternating single and multiple bonds, or lone pairs, creating at least one unhybridized p-orbital for each atom in the ring)
4. The compound has 4n+2 pi electrons (Huckel’s rule)

Question 14

Watson-Crick Model

Answer 14

Key features: the two strands of DNA are antiparallel or oriented in opposite directions

• the sugar-phosphate backbone is on the outside of the helix with the nitrogenous bases on the inside
• specific base pairing rules (complementary base pairing) A to T (or U) and G to C (C-G is three hydrogen bonds making it stronger
• %A=%T and %C=%G total purines are = to total pyrimidines Chargaff’s rules

Question 15

B-DNA

Answer 15

Right handed helix makes a turn every 3.4 nm and contains about 10 bases within that span
Most DNA

Question 16

Z-DNA

Answer 16

Zigzag appearance

Left handed helix turn every 4.6 nm and contains about 12 bases within each turn

Question 17

Denaturing of DNA

Answer 17

Disruption of hydrogen bonds
No covalent bonds between nucleotides in the backbone of the DNA break
Heat, alkaline pH, and chemicals like formaldehyde and urea cause this

Question 18

Renealing DNA

Answer 18

If denaturing condition is slowly removed two strands can become paired again.

Question 19

Probe DNA

Answer 19

DNA with a known sequence

Question 20

Histones

Answer 20

Small basic proteins around which the DNA that make up a chromosome are wound, creating a chromatin
There are two copies of H2A, H2B, H3, and H4 that form a histone core and about 200 base pairs of DNA are wrapped around the protein complex

Question 21

Nucleosome

Answer 21

Formed from the 200 base pairs of DNA that are wrapped around a histone core in chromosome

Question 22

Nucleoproteins

Answer 22

Proteins that associate with DNA

Histones fall into this category

Question 23

Heterochromatin

Answer 23

Small percentage of chromatin that remains compacted during interphase of cell cycle
Transcriptionally silent

Question 24

Euchromatin

Answer 24

Dispersed chromatin during interphase and has genetically active DNA

Question 25

Telomere

Answer 25

Repeating unit at the end of replication (TTAGGG). Some is lost each replication but can be replaced by the enzyme telomerase. However this accounts for limited replication Telomeres have high GC-content that creates strong strand attractions at the end of chromosomes to prevent unraveling

Question 26

Centromeres

Answer 26

Region of DNA found in center of chromosomes Composed of heterochromatin with repeating sequence and high GC content allowing two sister chromatids to remain connected until microtubules separate during anaphase

Question 27

Replisome

Answer 27

Also called replication complex - in DNA replication is a set of specialized proteins that assist the DNA polymerases

Question 28

Origins of replication

Answer 28

Point at which dna unwinds to begin replication

Question 29

Replication forks

Answer 29

Produced as DNA replication proceeds in both directions from the origins of replication

Question 30

Helicase

Answer 30

The enzyme responsible for unwinding DNA generating two single-stranded template strands ahead of the polymerase

Question 31

Single-stranded DNA binding proteins

Answer 31

Required to hold the separated strands of DNA apart during process of replication They bind to unraveled strand preventing both reassociation of the dna strands and the degradation of dna by nucleases

Question 32

DNA Topoisomerases

Answer 32

Introduce negative supercoils to relieve torsional stress of winding Work ahead of helicase by nicking one or both strands allowing relaxation of torsional pressure and then resealing the cuts.

Question 33

Parental strands

Answer 33

Serve as template for daughter strands

Question 34

Semi conservative

Answer 34

DNA replication is considered to be this because the parent strand is conserved in the two daughter strands

Question 35

DNA Polymerase

Answer 35

Responsible for reading the DNA template or parent strand and synthesizing the new daughter strand Can read template strand in a 3’ to 5’ prime direction while synthesizing complementary strand in the 5’ to 3’ direction

Question 36

Leading strand

Answer 36

Strand that is copied in a continuous fashion in the same direction as the advancing replication fork

Question 37

Lagging strand

Answer 37

Strand that is copied in a direction opposite the direction of the replication fork. DNA polymerase cannot synthesize directly because it only synthesizes in 3’ to 5’ direction.

Question 38

Okazaki fragment

Answer 38

Since DNA polymerase can only synthesize in 3’ to 5’ direction these fragments are developed for synthesis of the lagging strand

Question 39

Primase

Answer 39

This is before the replication begins, primase synthesizes a short primer of RNA (roughly 10 nucleotides) in the 5’ to 3’ direction for the DNA to hook onto as it splits Constantly laid down for lagging strand while leading strand requires only one

Question 40

DNA polymerase III

Answer 40

Prokaryotes only | Synthesizes daughter strand of DNA in the 5’ to 3’ manner

Question 41

DNA polymerases alpha, delta and epsilon

Answer 41

Eukaryotes only | Begins synthesizing daughter strands in the 5’ to 3’ manner

Question 42

Removal of RNA after synthesis in prokaryotes

Answer 42

DNA polymerase I

Question 43

Removal of RNA after DNA replication in eukaryotes

Answer 43

Carried out by RNase H

Question 44

Adds nucleotides where RNA primer had been, in prokaryotic dna replication

Answer 44

DNA polymerase

Question 45

Adds dna nucleotides where RNA primer had been in dna replication of eukaryotes

Answer 45

DNA polymerase delta

Question 46

DNA ligase

Answer 46

After DNA replication this seals the ends of DNA molecules together creating one continuous strand

Question 47

Five classic DNA polymerases in eukaryotes

Answer 47

Alpha, beta, gamma, delta, and epsilon Alpha, delta and epsilon work together to synthesize both the leading and lagging strands; delta also fills in gaps left behind when RNA primers are removed Gamma replicates mitochondrial dna Beta and epsilon- dna repair Delta and epsilon assisted by PCNA protein which assembles a trimer to form sliding clamp

Question 48

Sliding clamp

Answer 48

Clamp that helps strengthen interaction bt DNA polymerases delta and epsilon and the template strand

Question 49

Oncogenes

Answer 49

Mutated genes that cause cancer | Before mutation they are referred to as proto-oncogenes

Question 50

Antioncogenes

Answer 50

Tumor suppressor genes (p53 and Rb) | Encode proteins that inhibit the cell cycle or participate in dna repair processes

Question 51

Proofreading

Answer 51

In part of the polymerase - detects incorrectly paired bases during replication Distinguishes the template strand from daughter strand because template is more methylated Occurs during S cell cycle

Question 52

Mismatch repair

Answer 52

G2 phase is cell cycle Enzymes coded by genes MSH2 and MLH1 which detect and remove errors introduced in replication that were missed during the S phase of the cell cycle.

Question 53

Nucleotide excision repair

Answer 53

Eliminates thymine dimers introduced from UV light A cut and patch process Specific proteins scan the DNA molecule molecule and recognize the lesion An excision endonuclease makes nicks on both sides of thymine dimer and removes it Occurs during G1, G2 cell cycle

Question 54

Base excision repair

Answer 54

Detects small non-helix distorting mutations First affected base is recognized and removed by glycosylase enzyme, leaving behind an apurinic/apyrimidine (AP) site also called an abasic site AP site is then recognized by an AP endonuclease that removes damaged sequence and lets DNA polymerase and DNA ligase repair the gap Occurs during G1, G2 cell cycle

Question 55

Recombinant DNA

Answer 55

Allows dna fragment from any source to be multiplied by either gene cloning or polymerase chain reaction (PCR) which provides a means of analyzing and altering genes and proteins

Question 56

DNA cloning

Answer 56

Technique that can produce large amounts of a desired sequence DNA to be cloned is often present in small amounts and is part of a heterogeneous mixture of other DNA The goal is a large quantity of one type

Question 57

Vector or recombinant vector

Answer 57

In DNA cloning a piece of nucleic acid into which the dna is litigated by the investigator

Question 58

Restriction enzymes (restriction endonucleases)

Answer 58

Enzymes that recognize specific DNA sequences The 5’ to 3’ orientation of both strands is identical Isolated from bacteria Allows fragments to be inserted directly into the vector sometimes.

Question 59

DNA libraries

Answer 59

Large collections of known DNA sequences

Question 60

Genomic libraries

Answer 60

DNA library that contains large fragments of DNA and includes both coding (exon) and noncoding (intron) regions of genome

Question 61

cDNA (complementary DNA) library

Answer 61

DNA library constructed by reverse transcribing processes mRNA Lacks noncoding regions and only includes the genes that are expressed in the tissue from which the mRNA was isolated Sometimes called expression libraries

Question 62

Hybridization

Answer 62

Joining is complementary base pair sequences Can be DNA-DNA or DNA-RNA recognition Vital part of polymerase chain reaction and Southern blotting

Question 63

Polymerase chain reaction (PCR)

Answer 63

Automated process that can produce millions of copies of a DNA sequence without amplifying the DNA in bacteria Knowing sequences that flank the desired region of DNA allows for amplification of the sequence between

Question 64

Primers

Answer 64

PCR requires primers complementary to the DNA that flanks the region of interest, nucleotides (dATP, dTTP, dCTP, dGTP) and DNA polymerase Also needs heat Must use DNA polymerase from Thermus aquaticus - bacteria because ours cannot survive at high heat.

Question 65

Preferred gel for DNA electrophoresis

Answer 65

Agarose gel

Question 66

Southern blot

Answer 66

Used to detect presence and quantity of various DNA strands in a sample. DNA is cut by restriction enzymes and then separated by gel electrophoresis DNA fragments are then carefully transferred to a membrane, retaining separation Split strands bound to radioisotopes and indicates presence of desired sequence

Question 67

Dideoxyribonucleotide

Answer 67

A modified base that is added in lower concentrations in DNA sequencing Once added the polymerase can no longer add to the chain Include ddATP, ddCTP, ddGTP, ddTTP)

Question 68

Gene therapy

Answer 68

Potential cures for individuals with inherited diseases | Transferring a normal gene into the affected tissues

Question 69

Transgenic mice

Answer 69

Altered at their germ line by introducing a cloned gene into fertilized ova or into embryonic stem cells Cloned gene is called a transgene

Question 70

Knockout mice

Answer 70

Mice in which a gene has been intentionally deleted.

Question 71

Chimera

Answer 71

Blastocyst that has been given transgene but also has the original cells that lacks the transgene and has patches of cells