P1 Spring Pceutics 2/28 And 3/02

Front 1

Enzymes and chemicals used to make synthetic DNA from mRNA?

Back 1

1.Reverse transcriptase
2.Alkali treatement (separates mRNA from DNA strand)
3.DNA polymerase (makes complement of DNA strand)

Front 2

Tests that determine the quantity and quality of DNA?

Back 2

1.Spectrophotometric analysis
2.Restriction enzyme digests + Agarose gel electrophoresis

Front 3

What is the cloning of genes called that uses a gene fragment put into a plasmid, and transformation into bacteria?

Back 3

Subcloning

Front 4

Process that denatures a plasmid, adds a synthetic (mutated) oligonucleotide, extends it, and replicates it in E. coli (typically)

Back 4

Mutagenesis

Front 5

Steps in PCR?

Back 5

1.Separate strands with heat (94 deg) for 1 minute
2.Attach primers to single strands
3.Add DNA polymerase to extend strands
4.Repeat

Front 6

Purpose of Southern Blotting versus Western Blotting?

Back 6

Southern Blotting: to identify a specific gene (DNA) sequence

Western Blotting: to determine the relative amount of one protein from a sample of many

Front 7

Basic steps of Southern Blotting?

Back 7

1.Cut DNA (with RE)
2.Run on agarose gel
3.Denature fragments (helps fragments stick to membrane and probe)
4.Blot onto membrane
5.Add probe (a fluorescent DNA or RNA fragment, not an Ab)

Front 8

Mechanism of action for real time PCR?

Back 8

A probe that has a reporter and a quencher. When the reporter breaks off, it fluoresces.

Front 9

Start codons

Back 9

Methionine

ATG (DNA)

AUG (RNA)

Front 10

Stop Codons

Back 10

"You go away, you are away, you are gone"

UGA, UAA, UAG (RNA)

TGA, TAA, TAG (DNA)

Front 11

Antibiotic resistance genes added to plasmids when making recombinant DNA

Back 11

1.penicillin
2.ampicillin
3.kanamycin

Front 12

During DNA replication, new strand is made in what direction?

Back 12

5` to 3`

Front 13

What type of polymerase is used during transcription?

Back 13

RNA polymerase 2 (+ transcription factors)

Front 14

What does a DNA gene segment contain?

Back 14

•Sequence elements
•Promoter
•Start codon
•Exons/introns
•Stop codon
•Terminator
•Sequence elements

Front 15

What does a "transcription unit" contain?

Back 15

•Start codon
•Exons/introns
•Stop codon

Front 16

What ribosomal subunits exist during initiation and elongation?

Back 16

Initiation: 30s

Elongation: 70s (30s+50s)

Front 17

What enzyme attaches aminos to tRNA?

Back 17

aminoacyl synthetase

Front 18

Transformation versus transfection?

Back 18

Transformation: uptake of genetic material (eg plasmids) by bacteria

Transfection: uptake of genetic material (eg plasmids) by eukaryotes

Front 19

Common restriction enzyme sites?

Back 19

EcoRI, PstI, KpnI, XhoI

Front 20

Transformation enhanced by?

Back 20

Electric shock, chemicals

Front 21

LacZ gene codes for?

Back 21

B galactosidase (enzyme that breaks down lactose)

Front 22

LacZ gene used for?

Back 22

In recombination, codes for an identifier of our protein product:

B-gal + MET + OurProtein

Front 23

How is B-gal cleaved from our protein product?

Back 23

Cyanogen bromide

Front 24

Restriction enzymes that create single stranded overhangs (sticky ends)?

Back 24

Hind III (from Hemophilus Influenza)

Kpn I (from klebsiella pneumonia)

Front 25

Restriction enzymes that create blunt ends?

Back 25

Sma I

Front 26

Transfection enhanced by?

Back 26

electric shock, salts, lipids, dextrans

Front 27

Difference between bacterial and mammalian protein purification?

Back 27

Mammalian purification uses hollow core fibers