Chapter Ten: Digital Analysis of DNA

Question 1

___ fragment the genome at specific sites

Answer 1

restriction enzymes

Question 2

each restriction enzyme recognizes a ___

Answer 2

specific sequence of bases

Question 3

restriction enzymes cut ___

Answer 3

sugar-phosphate backbones of both strands

Question 4

___ are generated by digestion of DNA with restriction enzymes

Answer 4

restriction fragments

Question 5

___ is a commonly used restriction enzyme

Answer 5

EcoR1 from E. coli

Question 6

restriction enzyme cuts are straight through both strands of DNA at the line of symmetry

Answer 6

blunt ends

Question 7

restriction enzyme cuts are displaced on either side of the line of symmetry

Answer 7

sticky ends

Question 8

sticky ends have either 5’ or 3’ ___

Answer 8

overhangs

Question 9

sticky ends are good for when you want to make ___

Answer 9

recombinant DNA

Question 10

average restriction fragment size is ___

Answer 10

256 base pairs

Question 11

___ can be used to fragment DNA at random locations

Answer 11

mechanical forces

Question 12

mechanical forces can break ___

Answer 12

phosphodiester bonds

Question 13

two forms of mechanical forces

Answer 13

passing DNA through a thin needle at high pressure
sonication (ultrasound energy)

Question 14

___ distinguishes DNA fragments according to size

Answer 14

gel electrophoresis

Question 15

DNA has a ___ charge

Answer 15

negative

Question 16

___ fragments move further through gel

Answer 16

small

Question 17

a means to purify a specific DNA fragment away from all other fragments and make many copies of the fragment

Answer 17

molecular cloning

Question 18

two basic steps of molecular cloning

Answer 18

1. insert DNA fragments into coning vectors to specialized chromosome-like carriers that ensure transport, replication, and purification of DNA inserts
2. transport recombinant DNA into living cells to be copied

Question 19

a group of replicated DNA molecules

Answer 19

DNA clone

Question 20

three main features of a plasmid cloning vector

Answer 20

1. origin of replication
2. selectable marker gene
3. synthetic polylinker

Question 21

DNA sequence containing multiple restriction enzyme sites

Answer 21

synthetic polylinker

Question 22

___ and ___ are alternate cloning vectors that can carry large inserts

Answer 22

bacterial artificial chromosomes (BAC) and yeast artificial chromosomes (YAC)

Question 23

create ___ using plasmid vectors

Answer 23

recombinant DNA

Question 24

digestion of the vector and human genomic DNA with a ___ results in ___

Answer 24

restriction enzyme
complementary sticky ends

Question 25

___ is used to deal the phosphodiester backbones between vector and inserted fragment

Answer 25

DNA ligase

Question 26

process by which a cell or organism takes up foreign DNA

Answer 26

transformation

Question 27

only cells that ___ will grow on media specific to the selectable marker gene included in the plasma

Answer 27

took up the plasmid

Question 28

long-lived collection of cellular clones that contains copies of every sequence in the whole genome inserted into a suitable vector

Answer 28

genomic library

Question 29

a perfect genomic library has one copy of ___

Answer 29

every sequence in the entire genome

Question 30

___ to obtain a perfect library

Answer 30

impossible

Question 31

Sanger sequencing uses ___ to make new DNA

Answer 31

DNA polymerase

Question 32

DNA polymerase requires these 4 things

Answer 32

1. template strand to copy
2. deoxyribonucleotide triphosphates
3. primer
4. buffer

Question 33

a ___ is a good template for Sanger sequencing

Answer 33

recombinant plasmid

Question 34

cloned recombinant DNA is ___

Answer 34

denatured

Question 35

strands are mixed with ___

Answer 35

oligonucleotide primer

Question 36

as temperature is lowered, primers and template strand ___

Answer 36

anneal (hybridize)

Question 37

hybridized template and primer are mixed with ___, ___, and ___

Answer 37

DNA polymerase, dNTPs, and small amounts of dideoxyribonucleotide triphosphates (ddNTPs)

Question 38

ddNTPs each have a unique ___

Answer 38

fluorescent tag

Question 39

ddNTPs lack a ___, so once they are added to the new DNA strand ___

Answer 39

3’ OH
polymerization stops

Question 40

result of this is a set of ___ with different 3’ ends

Answer 40

nested fragments

Question 41

nested fragments are separated by size using ___

Answer 41

electrophoresis

Question 42

___ is recorded to identify nucleotide code

Answer 42

color of terminal ddNTP

Question 43

E. coli have ___ chromosome, ___ genes, and a genome size of ___

Answer 43

1, 4400, 4.6 Mb

Question 44

yeast have ___ chromosomes, ___ genes, and a genome size of ___

Answer 44

16, 6000, 12.5 Mb

Question 45

humans have ___ chromosomes, ___ genes, and a genome size of ___

Answer 45

23, 2800, 3300 Mb

Question 46

the human genome project began using the ___

Answer 46

hierarchical strategy

Question 47

steps of hierarchical strategy of sequencing genomes

Answer 47

1. construct BAC genomic library
2. identify sets of overlapping BAC clones
3. shear DNA from each BAC separately to make smaller clones
4. sequence DNA based on overlap

Question 48

steps of whole-genome shotgun strategy for genome sequencing

Answer 48

1. create genomic library of overlapping fragments in plasmid vectors
2. sequence DNA inserts of randomly chosen library plasmids
3. assemble sequences based on overlap of sequences into contigs

Question 49

the whole genome shotgun approach can be highly ___

Answer 49

automated

Question 50

sequencing a BAC clone library rather than smaller inserts of plasma clones

Answer 50

paired-end sequencing

Question 51

trying to figure out what genes code for

Answer 51

annotation