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1st Year sem 2: Forensic Identification > V L8: DNA amplification and visualisation > Flashcards

V L8: DNA amplification and visualisation Flashcards

1
Q

where are restriction enzymes found?

A

mainly in bacteria

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2
Q

whats the full name for restriction enzyme?

A

Bacterial endonuclease enzymes

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3
Q

Bacterial endonuclease enzymes are used to ___ up human ____.

A

snip

DNA

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4
Q

what do restriction enzymes recognise?

A

recognise restriction site which is 4-8 base pairs

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5
Q

restriction enzymes recognise sites that are _____.

A

palindromic

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6
Q

palindromic is read same in every direction, true or false?

A

true

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7
Q

what are the two types of cut?

A

staggered cuts

straight cut

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8
Q

A staggered cuts leaves what ends?

A

sticky ends

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9
Q

A straight cut leaves what end?

A

blunt ends

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10
Q

How do Restriction Fragment Length Polymorphisms appear?

A
  • a mutation at the particular restriction enzyme site
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11
Q

Restriction Fragment Length Polymorphisms shows as?

A

only 1 fragment is labelled, the second fragment is larger.

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12
Q

what causes DNA separation?

A

size

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13
Q

larger fragments travel _____.

A

slower

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14
Q

DNA sample is mixed with a ____ ______, this leaves cut up _____.

A

restriction enzyme

DNA

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15
Q

when was the first time DNA was sued in a double murder case?

A

Colin pitchfork, 1980

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16
Q

what was the technique previously step by step?

A 
extraction
chopping up 
separating (electrophoreses)
southern blotting 
DNA immobilised 
xray film
DNA fingerprint
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17
Q

DNA fingerprinting is an ____ technique.

A

older

18
Q

Minisatellite DNA is?

A

VNTRs

19
Q

VNTRs produce a ____ portion fo DNA

A

larger

20
Q

VNTRs are more ____.

A

variable

21
Q

whats used now VNTRs or STR?

A

STR

22
Q

STR

A

target certain portion of genome, will either have same or different number of repeats on chromosomes,

23
Q

PCR is?

A

polymerase chain reaction

24
Q

Microsatellite DNA is also known as?

A

short tandem repeats (STR)

25
Q

STRs are prone to _____ therefore are useful for ____ _______.

A

mutation

genetic profiling

26
Q

PCR was developed in?

A

1985

27
Q

PCR allows us too?

A

You can amplify DNA form a very small amount

28
Q

what are the 3 main stages of PCR?

A

Denaturation (heat) - form single strand
Annealing (hybridisation of primers) - primers bind to specific area
Extension - DNA polymerase and building blocks produce new strand (extend primers)

29
Q

with every cycle of PCR we ____ the amount of DNA.

A

double

30
Q

why are STRS used more now?

A
  • Due to the amount of DNA required to perform the southern blot procedure
  • large PCR amplicon size of VNTR’s
31
Q

what is the DNA amplification cycle?

A
  1. Denaturation at high temperature (92°C-95°C)
  2. Annealing at a cooler annealing temperature
  3. Extension at a temperature between the annealing and denaturing temperatures (usually at 72°C)
32
Q

annealing temperature depends on?

A

primers the sequence or length of them

33
Q

Tm is..

A

melting temperature

34
Q

extension dependings on?

A

the DNA polymerase

35
Q

after PCR DNA should be form this equation?

A

DNA=2N

36
Q

what are the PCR compoennts?

A
  • Template DNA (0.5-2.5 ng)

- Reagents, DNA polymerase, Oligonucleotide primers, buffer, DNTP

37
Q

DNTP are the ?

A

building blocks

38
Q

most enzymes need what to work?

A

ions

39
Q

DNA polymerase needs a co factor such as?

A

magnesium ions

40
Q

when do we produce fragments of the right size?

A

third cycle on wards

41
Q

what to look for in a PCR primer?

A
  • Conserved regions of DNA
  • Complementary to the flanking sequence of the target region
  • similar melting temperature for forward and reverse primer
  • 18-30 bases long
  • should NOT contain self-complementary sequences

1st Year sem 2: Forensic Identification (15 decks)

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