Lecture 9/14 - E2 Flashcards

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1
Q

Common Temperature for Denaturation

A

94-98C

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2
Q

Common Temperature for Annealing

A

37-60C

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3
Q

Common Temperature for Extension

A

68-72C

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4
Q

How do you calculate the total number of amplified products after each cycle?

A

2^n

n = the number of cycles

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5
Q

How do you calculate the total number of ds DNA products when you begin with a single ds DNA molecule?

A

A(2^n - 2n)
A = the number of starting ds DNA template molecules
n = number of cycles

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6
Q

Advantages of PCR over traditional approaches?

A

Faster and less expensive, can be optimized to detect low abundance mRNA splice variants, and it can be used for other purposes

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7
Q

Limitations of PCR technology

A

Requires some precise knowledge of target DNA sequence in order to create the primers, size restrictions on amplicons, mutations may be introduced into the DNA by the polymerase during amplification

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8
Q

Starting with genomic dsDNA isolated from a single somatic cell, how many fully dsDNA products are present after 4 cycles of PCR?
A) 0
B) 2
C) 4
D) 6
E) Cannot determine based on the information providded

A

E

Need to know how many molecules are present at the start of the reaction which depends upon the stage of the cell cycle

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9
Q
Starting with genomic dsDNA from a single somatic cell at the G1 stage of the cell cycle, how many fully dsDNA products are present after 4 cycles of PCR?
A) 0
B) 4
C) 8
D) 16
E) 32
A

There are 2 tmplates because of mom and dad copies
A(2^n - 2n)
2(2^4 - 2(4)) = 2(16-8) = 16

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10
Q

Haplosufficient

A

Where one copy of the WT allele is needed for the full WT functionality (Aa)

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11
Q

Haploinsufficient

A

Where one copy of the fully functioning allele shows a mutant phenotype (Aa = mutant)

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12
Q

Null/Amorphic Mutations

A

Complete loss of function mutation that produces no functional gene product
Completely inactivate gene function

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13
Q

Weak/Leaky/Hypomorphic Mutations

A

Mutations that result in partial loss / reduced function

- Depends on the level of activity of the leaky mutant allele

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14
Q

Possible defects associated with Null Alleles

A
Gene absent or inactivated
No transcription
Abberrant proessing of the mRNA transcript
No translation
Non-functional protein product
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15
Q

Possible defects associated with Hypomorphic Alleles

A
No gene
Reduced rate of transcription
Abberant processing of the mRNA transcript
Reduced translation
Less-functional protein product
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16
Q

Which types of DNA change in the yeast ACT1 gene is most likely to result in a hypomorphic mutation?
A) Deletion of all ACT1 protein coding sequence
B) Removal of all teanscriptional regulatory sequences
C) A codon change resulting in an amino acid substitution
D) ACT1 gene duplication
E) All equally likely to cause hypomorphic mutation

A

C) A codon change resulting in an amino acid substitution

Because it is the least mild change

17
Q

Hypermorphic Allele

A

Increased activity (more is not always better)

18
Q

Neomorphic Mutation

A

Mutation that results in an increased function

19
Q

Dominant Negative Mutation

A

Competes with the WT protein and blocks natural function
Produces poisonous gene products
“Spoiler Effect” - Homo dominant can be lethal and too toxic

20
Q
Which type of mutation is most likely to result in a haploinsuficient genotype?
A) A null allele
B) A weak hypomorphic allele
C) A wildtype allele
D) A gene duplication
A

A) A null allele