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Forensic Biology > DNA Extraction > Flashcards

DNA Extraction Flashcards

1
Q

As of 2020, how many people have the Innocence Project exonerated using DNA?

A

375

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2
Q

1989

A

the first DNA exoneration took place

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3
Q

how many exonerated cases started with eyewitness misidentification?

A

69%

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4
Q

reminders about DNA collection/storage

A
  • dry out samples to limit contamination from mold
  • breathable container
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5
Q

how to collect samples (dry blood in tub)

A
  • scrape dry blood into an envelope
  • wet swab to get blood then let it dry
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6
Q

collecting reference/known samples

A
  • fast
  • painless
  • high DNA yield
  • buccal swabs
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7
Q

preferred methods of sample collections

A

buccal swabs

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8
Q

sample storage

A

FTA cards

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9
Q

samples inside of the lab

A
  • DNA samples are extracted & either stored in a refrigerator at 4°C or a freezer at -20°C
  • for long term storage, extracted DNA samples may be stored at -70°C
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10
Q

DNA Extraction: what does the soap solution do?

A

breaks open cells by disrupting the phospholipid bilayer

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11
Q

DNA Extraction: what does the saline solution do?

A

helps DNA stick together by separating macromolecules from DNA

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12
Q

DNA Extraction: what does the alcohol do?

A

clumps DNA together so it can be removed

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13
Q

Chelex Extraction

A
  • add Chelex 100 suspension to sample & mix
  • heat sample at 95°C for 2-10 minutes
  • separate Chelex 100 beads from sample by centrifugation, setting, or filtration
  • use supernatant in downstream step such as PCR, RT-qPCR, RT-ddPCR, or RT-LAMP
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14
Q

DNA Quantification- Why?

A
  • cost effective to screen for DNA first (could be none present)
  • need to determine before PCR because a narrow range of DNA is optimal for most STR typing
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15
Q

Real Time Quantification PCR (RT-qPCR)

A
  • determines the amount of amplifiable DNA
  • reflects both quantity and quality
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16
Q

RT-qPCR analysis

A
  • cycle to cycle change in fluorescence
  • more DNA = more fluorescence
17
Q

steps of Polymerase Chain Reaction (PCR)

A
  • denaturing
  • annealing
  • extending
  • repeat
18
Q

PCR: denaturing

A

expose DNA to raised temperature & the H2 bonds between base pairs break & the 2 strands of the DNA molecules separate

19
Q

PCR: annealing

A

lowering temperature & primers will attach through complimentary base pairing

20
Q

PCR: extending

A
  • raise the temperature
  • DNA polymerase extends the new strand of DNA from the primers using dNTPs
21
Q

CT - cycle threshold

A

number of PCR cycles for fluorescence to signal threshold

22
Q

qPCR CT values inverse relationship

A

inverse relationship between CT values & amount of DNA in the sample

23
Q

after 1 round of PCR, 1 molecule of DNA consisting of 2 complementary strands yields ___ molecules of DNA, for a total of ___ strands

A

2, 4

24
Q

how many DNA molecules would exist after 10 PCR cycles?

A

10^2 = 100 DNA molecules

25
Q

how many DNA molecules would exist after 30 PCR cycles?

A

30^2 = 900 DNA molecules

26
Q

PCR

A
  • allowing testing of partially degraded & small samples
  • shortened analysis times from weeks to days
  • increased ability to use DNA evidence
27
Q

who developed PCR?

A

Kary Mullis

28
Q

why is PCR not the whole answer?

A

it can only duplicate smaller fragments

29
Q

short tandem regions

A

the repeat region is variable between samples while the flanking regions where PCR primers bind are constant

30
Q

homozygote

A

both alleles are the same length

31
Q

heterozygote

A

alleles differ & can be resolved from one another

32
Q

calculate: “AAGCTA” DNA fragment length: 72 bp # of repeats

A

72/6 = 12

33
Q

calculate: “TAGGG” DNA fragment length: 105 bp # of repeats

A

105/5 = 21

34
Q

where are STR’s compared?

A

multiple sites

Forensic Biology (17 decks)

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