Chapter 17

Question 1

Gel electrophoresis

Answer 1

• Separates DNA fragments by size
• Fragments usually produced using restriction endonucleases
• Gel (agarose or polyacrylamide) is submersed in a buffer carrying electrical current
• Negatively-charged DNA migrates toward positive pole
• Large fragments move slowly, small move fast
• DNA is visualized using fluorescent dyes

Question 2

Polymerase chain reaction (PCR)

Answer 2

Amplifies small DNA fragments using primers that flank the region

Question 3

Each PCR cycle involves three steps:

Answer 3

• Denaturation (high temperature)
• Annealing of primers (low temperature)
• DNA synthesis (intermediate temperature). Uses Taq polymerase (a DNA polymerase from Thermus aquaticus); can tolerate relatively high temperatures.

Question 4

Application of PCR

Answer 4

• Allows the investigation of minute DNA samples
• Forensics: drop of blood, cells of a hair follicle
• Detection of genetic defects in embryos by analyzing a single cell
• Analysis of mitochondrial DNA

Question 5

PCR components

Answer 5

• Primers: short pieces of DNA complementary to each end of the target DNA fragment
• Tag polymerase
• Deoxyribose nucleotide triphosphates (ATP, CTP, GTP, TTP)

Question 6

Southern blotting

Answer 6

• IDs a known DNA sequence in an unknown DNA sample
• DNA fragments from an unknown are separated using gel electrophoresis, and then transferred to a membrane (blotted)
• The blot is flooded with a labelled DNA probe with a base sequence complementary to a known (target) sequence
• The probe binds to fragments in the unknown sample if the target base sequence is present

Question 7

clone

Answer 7

genetically identical copy

Question 8

molecular cloning

Answer 8

multiplication of a specific DNA sequence. Sometimes called gene cloning

Question 9

vector

Answer 9

carries DNA into the host and can replicate in the host

Question 10

plasmids

Answer 10

small circular DNA molecules

Question 11

viruses

Answer 11

can insert DNA into cells

Question 12

Types of vectors:

Answer 12

• plasmids
• viruses
• yeast artificial chromosomes (YACs)
• bacterial artificial chromosomes (BACs)

Question 13

YACs and BACs

Answer 13

allow for larger insert of large-scale analysis of genomes

Question 14

plasmid vectors

Answer 14

small, circles of DNA used to clone small pieces of DNA

Question 15

Selectable marker gene

Answer 15

allows presence of plasmid to be identified in host cells (often an antibiotic resistance gene)

Question 16

multiple cloning site (MCS)

Answer 16

area with many different restriction sites that allow the plasmid to be cut at many different points

Question 17

bacteriophage (viruses that infect bacteria)

Answer 17

can also be used to move DNA into cells

Question 18

DNA libraries

Answer 18

A collection of DNAs in host cells that, when considered together, represent the complete DNA sequence

Question 19

Genomic library

Answer 19

represents the entire genome in a set of host cells. DNA is fragmented, inserted into vectors (e.g., BACs), and reproduced in host cells.

Question 20

cDNA

Answer 20

DNA copies of mRNA are made using the enzyme reverse transcriptase

Question 21

mRNA extracted from tissue

Answer 21

represents only expressed genes, no introns

Question 22

cDNA is used to make and store the

Answer 22

cDNA library

Question 23

Genomic libraries from a single organism will be the … but cDNA libraries will …

Answer 23

same, differ

Question 24

Dolly the Sheep

Answer 24

• Dolly the sheep was the first cloned mammal.
• The nucleus was removed from a donor egg cell.
• The nucleus from a second sheep was then introduced into the cell, which was allowed to divide before implanting in a surrogate mother.

Question 25

Medically important proteins:

Answer 25

- Human insulin, interferon, atrial peptides - Tissue plasminogen activator - Human growth hormone

Question 26

Vaccines:

Answer 26

- Subunit vaccines; Genes encoding a part of a viral protein are spliced into part of the vaccinia (cowpox) genome - Injection of harmless recombinant virus leads to immunity

Question 27

gene therapy

Answer 27

- adding a good copy of a gene to correct genetic disorder

Question 28

Severe combined immunodeficiency disease (SCID) illustrates both the potential and the problems of gene therapy

Answer 28

- Positive: 15 children treated successfully - Negative: 3 other patients developed leukemia due to therapy

Question 29

Transgenic (GMO) Animals

Answer 29

contains recombinant DNA used to express human genes Ex. mice, sheep, goats, mosquitoes

Question 30

Transgenic (GMO) Plants

Answer 30

Contain genes for resistance to disease, herbicides, and/or pesticides Ex. Bt corn, Flavr Savr Tomato, Transgenic Chestnut

Question 31

Bt corn

Answer 31

Contains Bt gene, which produces a toxin that kills insects. Presumably non-toxic to humans and other mammals.

Question 32

Flavr Savr Tomato

Answer 32

has genes that slow softening and fungal rot

Question 33

Transgenic Chestnut

Answer 33

has gene from wheat that controls chestnut blight

Question 34

CRISPR/Cas9

Answer 34

- can be “programmed” to cut DNA at any specified location (base sequence) - Can delete particular genes, or can be used to cut out and replace existing genes with other genes (gene editing)

Question 35

Predicting Disease Risk

Answer 35

Changes in transcription often represent the earliest response to disease

Question 36

PCA3

Answer 36

is expressed in prostate epithelial cells, but overexpressed in cancerous cells

Question 37

BRCA1

Answer 37

normally helps repair damaged DNA. Mutations in BRCA1 are associated with breast cancer.

Question 38

PCA3 and BRCA1 are examples of

Answer 38

molecular markers of disease

Question 39

genomics

Answer 39

study of entire genomes at level of DNA

Question 40

proteomics

Answer 40

study of entire set of proteins

Question 41

metabolomics

Answer 41

study of while set of small molecules found in an organism

Question 42

transcriptomics

Answer 42

study of entire set of mRNAs expressed in an organism

Question 43

mitochondrial genomics

Answer 43

maternally inherited to next generation

Question 44

metagenomics

Answer 44

study of many genomes in an environmental sample (soil, water)

Question 45

Pharmcogenomics (aka: Toxicogenomics)

Answer 45

study of transcription changes in response to a chemical/drug