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BMSC 210 > Module 7 > Flashcards

Module 7 Flashcards

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1
Q

What is heterologous expression?

A

Expressing a gene in a different host

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2
Q

What is genetic engineering

A

Using in vitro techniques to alter genes in the lab

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3
Q

What is DNA amplification

A

DNA replication in a test tube

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4
Q

How to the cycle numbers grow in PCR ?

A

Each future cycle doubles the number of copies.

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5
Q

What are the PCR steps?

A

I. Denature DNA
2, annealing → oligonucleotides added
3. Add DNA polymerase
4. Heat and cool

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6
Q

What is reverse transcription PCR

A

Makes DNA copies from mRNA template

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7
Q

What does gel electrophoresis do?

A

Separates DNA molecules based on size and charge

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8
Q

What is DNA’s charge?

A

Negative

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9
Q

What is nuclei acid probing

A

Used to detect a certain nucleic acid sequence

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10
Q

How is the probe usually labelled or tagged?

A

With fluorescence

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11
Q

What is a probe?

A

Probes are complementary to the gene of interest

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12
Q

What is a southern blot

A

DNA is in the gol and probe is RNA or DNA

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13
Q

What is a northern blot

A

RNA is in the gel and the probe is DNA or RNA

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14
Q

What is fluorescent in situ hybridization

A

Uses a fluorescent probe attached to oligonucleotide to target specific nucleic acid sequences in cells

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15
Q

What are the enzymes used for cloning?

A

Restriction endonucleases
DNA ligase
Reverse transcription
DNA polymerase

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16
Q

What are restriction endonucleases

A

Enzymes that recognize specific DNA sequences and cut them

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17
Q

What are restriction endonucleases essential for?

A

In vitro DNA manipulation and gene cloning

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18
Q

What is methylation?

A

Coding on cells to prevent cutting by restriction enzymes (defense)

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19
Q

Are restriction endonucleases rare in eukaryotes

A

Yes

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20
Q

What do type 2 restriction endonucleases do?

A

Cleave DNA within recognition sequence: most useful for specific DNA manipulation

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21
Q

What recognizes palindromes

A

Type 2 restriction endonucleases

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22
Q

What is also known as an overhang cut

A

Sticky end cut

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23
Q

What is blunt end cutting

A

Both strands are cut at the same position

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24
Q

When are sticky ends produced?

A

When restriction enzymes make staggered cuts

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25
What can molecules with complimentary sticky ends easily do?
Anneal or form hydrogen bonds between complimentary bases
26
What is ecoR1
A type 2 restriction enzyme that is isolated from strains of E coli
27
What is each restriction enzyme paired with?
A corresponding modification enzyme that shares the same recognition sequence
28
How can a gene to be cloned be amplified by? (3)
Polymerase chain reaction Synthesized by reverse transcriptase Synthetic DNA made in vitro
29
What are used as cloning vectors
Plasmids
30
What are the 3 main steps of gene clothing?
Isolation and fragmentation of source DNA Insertion of DNA fragment into cloning vector Transformation of cloned DNA into host organism
31
What is blue / white screening
Bacteria with a cloning vector inside that may or may not have recombinant DNA
32
What do blue colonies not have
Plasma vectors with foreign DNA inserted
33
What do white colonies have
The recombinant DNA
34
What gene does blue/ white screening rely on?
LacZ gene
35
Where does the lacZ gene often fuse?
To a portion of the plasmid known as the multiple cloning site
36
When X gal is cleaved by b-galactosidase, what happens?
It turns blue
37
Why does a colony stay white?
Because it contains the gene or interest and lacZ gene will be disrupted and b-galactosidase cannot be produced
38
What is on the plasmid vector
LacZ gene and a multiple cloning site
39
What are DNA cassettes / cartridges
Synthetic fragments that can make more then a few base pair changes or replace sections of a gene via cassette mutagenesis
40
what are the properties of an open reading frame (4)
ribosome binding site seuqunce before the start codon start codon coding sequence stop codon
41
what are the enzymes in DNA cloning
DNA polymerase Restriction endonuclease DNA ligase Reverse transcriptase
42
whats the purpose of DNA polymerase in cloning
it synthesizes the DNA strand
43
what is the purpose of restriction endonuclease in DNA cloning
they recognize specific DNA sequences and cut DNA
44
what is the purpose of DNA ligase in cloning
going two strands of DNA
45
what is the purpose of reverse transcriptase
converts RNA into DNA
46
what do expression vectors have that coning vectors don't
inducible promoter tag for purification
47
what are expression vectors for?
they are designed to facilitate gene expression and protein production
48
what is an inducible promotor?
a DNA sequence that drives the expression of the gene - it controls initiation of transcription by RNA polymerase
49
what is a cloning vector?
DNA molecule used to carry and replicate foreign DNA fragments in a host organism
50
what is the best generic method to diagnose an RNA virus
RT-PCR
51
why is the RT-PCR the best genetic method to diagnose an RNA virus
the reverse transcriptase allows the synthesis of cDNA from RNA
52
what is genomics
the study of the entire genome (all genes and genetic material)
53
what us transcriptomics
the study of a transcriptome, which is the complete set of RNA molecules (mainly mRNA)
54
what is proteomics
the study of the proteome, which is the entire set of proteins expressed by the genome
55
what is metabolomics?
the study if a metabolome, which is the small molecule metabolites (sugars, lipids, amino acids)
56
what does transcriptomics focus on understanding
gene expression patterns
56
what are microarrays
they use pre designed probes to measure the expression levels of thousands of genes at once
57
what is western blotting used for
measuring protein levels
58
why are primers required for polymerase chain reactions
to provide a starting point for DNA polymerase
59
what is a thermocycler
a machine used for PCR reactions
60
what is the structure of a nucleic acid probe
ss DNA fragments complementary to the sequence of interest
61
cutting DNA with EcoR1 will produce
5' overhangs

BMSC 210 (16 decks)

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