Final

Question 1

Transformation

Answer 1

DNA transfer into bacteria

Question 2

Transduction

Answer 2

Viral DNA transfer into bacteria

Question 3

Transfection

Answer 3

DNA transfer into eukaryote

Question 4

Plasmid DNA Isolation Steps

Answer 4

CPR ANCER

Culture
Pellet
Resuspend (Sln 1)
Alkaline Lysis (Sln 2)
Neutralization (Sln 3)
Centrifuge
Ethanol
Resuspend

Question 5

Alkaline Lysis Sln 1

Answer 5

GET

Glucose: Osmotically stable
EDTA: Chetlates Divalent Cations (Ca2+, Mg2+) and destabilizes membrane
Tris-Cl: Alkaline pH

Question 6

Alkaline Lysis Sln 2

Answer 6

SN

SDS: Detergent distrupts membranes and causes lysis, as well as denaturing proteins
NaOH: Increases pH to 12.5 to denature DNA, RNA, and Protein

Question 7

Alkaline Lysis Sln 3

Answer 7

AP

Acetic Acid: Neutralizes NaOH
Potassium Acetate: Potassium SDS is insoluble and precipitates with denatured proteins

Question 8

Methods to Analyze Plasmid DNA Prep

Answer 8

Agarose Gel Electrophoresis
Spectrophotometry

Question 9

Pure DNA A260/A270

Answer 9

About 1.8

Question 10

Pure RNA A260/A270

Answer 10

> = 2.0

Question 11

Conjugation

Answer 11

Interbacterial DNA transfer

Question 12

Chemical Methods

Answer 12

Heat Shock / CaCl2
Calcium Phosphate Incubation
Lipofection

Question 13

Physical Methods

Answer 13

Electroporation
Biolistics
Microinjection

Question 14

Heat-shock / CaCl2

Answer 14

Add DNA to cells chilled on ice
Heat-shock cells at 42C for 0.5 to 2 minutes

Chemical transformation
Inexpensive but inefficient

Question 15

DNA Calcium Phosphate Incubation

Answer 15

Chemical transfection
No heat-shock required, suitable for eukaryotic cells

Question 16

Lipofection

Answer 16

Negatively charge DNA mixed with cationic lipids to form liposomes

Question 17

Biolistics

Answer 17

DNA is adsorbed on gold microparticles and shot into cells using gas-pressurized gene gun

Question 17

Electroporation

Answer 17

Brief high-voltage electrical fields increase membrane permeability
10X more effective than chemical transformation

Question 18

Microinjection

Answer 18

Glass micropipette injects liquid solution or substance directly into cell

Question 19

Viral Transduction

Answer 19

Virus mediated transfer of DNA into cells

Question 20

X-gal

Answer 20

Lactose analogue that produces a blue colour when cleaved

Question 21

IPTG

Answer 21

Inducer of lac operon

Question 22

Reverse Transcriptase

Answer 22

Creates single stranded DNA from mRNA

Question 23

DNA Polymerase

Answer 23

Creased second strand of DNA to create double stranded cDNA

Question 24

Colony Hybridization

Answer 24

Filter is exposed to film, autoradiography is compared to original agar plate.

Question 25

PCR Stages

Answer 25

Denaturation at 95C Annealing at 65C Extension at 75C

Question 26

Bacteriophage Vectors

Answer 26

Efficient and commonly used for delivering DNA to bacterias

Question 27

Cosmids

Answer 27

Hybrid plasmid derived from lambda phage, can accomodate large DNA fragments

Question 28

Expression Vectors

Answer 28

High level of protein expression due to promoter site located next to MCS

Question 29

Shuttle Vectors

Answer 29

Modified plasmid that can replicate in either prokaryotic or eukaryotic cells.

Question 30

Sanger Sequencing

Answer 30

Like PCR, but with fluorescently labeled ddNTP that are missing hydroxyl group. Once strands of every size exist, fluorescent ddNTPs can be read in order to get sequence

Question 31

Prokaryote Promoter Regions

Answer 31

-10: TATAAT -35: TTGACA

Question 32

Prokaryote Translation Initialization

Answer 32

Shine-Dalgarno Sequence

Question 33

Eukaryote Translation Initialization

Answer 33

Kozac Consensus Sequence

Question 34

Transcription Termination Sequence in Prokaryote

Answer 34

Rho-dependent Rho-independent

Question 35

Eukaryote Termination Sequence

Answer 35

Involves polyA signal

Question 36

Polyadenylation in Eukaryotes

Answer 36

Invovled in nuclear export, translation, stability

Question 37

Polyadenylation in Prokaryotes

Answer 37

Promotes mRNA degredation

Question 38

Yeast Expression System Suitable for

Answer 38

Disulfide bonds and glycosylation

Question 39

Fungal Expression Systems used in

Answer 39

Alcohjol, industrial enzymes, citric acid, medication