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Applied Genetics > Restriction enzymes and DNA cutting > Flashcards

Restriction enzymes and DNA cutting Flashcards

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Biology 101 flashcards
1
Q

how is polyploidy genetic variation induced

A

need to add a chemical

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2
Q

what is genetic engineering

A

manipulation of DNA of organisms for the purposes of, changing inheritance or the in vitro manufacture of gene products

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3
Q

what is a mutation

A

any change in genetic information relative to reference ‘wild type’ genome

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4
Q

what is a mutagen

A

any agent causing mutations

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5
Q

how do mutations with mutagens occur

A

randomly

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6
Q

what occurs in genetic engineering

A

enable us to take genes out
genetic material analysed
DNA can be altered
DNA can be re-inserted in organism

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7
Q

how can DNA be cut

A

shaking DNA violently, but this will cause random breaking

DNA can be cut using enzymes in more specific and targeted way

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8
Q

what are the two enzyme types

A

Endonucleases

Exonucleases

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9
Q

what is an enzyme

A

a biological catalyst; all biochemical reactions are controlled by enzymes and in general, each enzyme controls each separate reaction

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10
Q

what is a restriction enzyme

A

alternative name for endonuclease

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11
Q

what is an exonuclease

A

cleaves nucleotides one at a time from the end (exo) of an polynucleotide chain

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12
Q

what did Werner Arber discover

A

restriction enzymes

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13
Q

what did Daniel Nathans discover

A

Pioneered the application of restriction for the construction of genetic maps

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14
Q

what did Hamilton Smith discover

A

Showed that restriction enzyme cuts DNA in the middle of a specific sequence

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15
Q

what happened in 1978

A

Arber, Nathans and Smith won a Nobel Prize

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16
Q

how was HindlI discovered

A

is the first restriction enzyme discovered accidentally in 1970; while studying how the bacterium Haemophilus influenzae defends itself to the virus

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17
Q

what can the bacteria Haemophilus influenzae do

A

break down the DNA from the phage and any other foreign DNA; yet its own DNA is not damaged

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18
Q

what does base Y mean

A

C+T

19
Q

what does base R mean

A

G+A

20
Q

what can exonucleases be used for

A
  • Removal of oligonucleotides before a PCR reaction
  • Removal of chromsomal DNA in plasmid preparations
  • Removal of DNA in RNA preparations
  • Generation of single strand DNA (ssDNA) from linear double strand DNA (dsDNA)
21
Q

what is recombinant DNA technology

A

creation of a new combination of DNA segments that are not found together naturally

22
Q

what are the applications of recombinant DNA technology

A

Basic research on control of gene expression
Forensic medicine
Biotechnology - generate new products

23
Q

how do restriction endonucleases work

A

Cleave phosphodiester bonds of nucleic acids at an internal site
highly specific; can recognise 4 bases pair (HaeIII), 6bp(EcoRI), 8bp (NotI) or more
restriction sites that are recognised are usually palindromic

24
Q

what is the use of a restriction endonuclease

A

Cut DNA internally and in a more specific and targeted way; their role is vital for the use of Genetic Engineering

25
Q

what is a palindrome

A

a sequence of units that can be read the same way in either direction

26
Q

what is the frequency of enzyme recognition

A

4n

n = number of bases in recognition sequence

27
Q

if each position of the recognition site has 4 possible bases what is the probability of finding any one base

A

1/4

28
Q

where does Haelll recognise a segment

A

segment of 4 bases
5’ GC/CC 3’
3’ CC/GG 5’

29
Q

where does Smal recognise a segment

A

segment of 6 bases
5’ CCC/GGG 3’
3’ GGG/CCC 5’

30
Q

where does Notl recognise a segment

A

segment of 8 bases
5’ GC/GGCCGC 3’
3’ CGCCGG/CG 5’

31
Q

what are blunt ends

A

end of a DNA molecule where both stands ends in a base pair

32
Q

what is an overhang

A

stretch of unpaired nucleotides in the end of a DNA molecule, they can be in either strand

33
Q

what are sticky ends

A

overhangs of more than one nucleotide at the end of a DNA molecule; also called cohesive ends

34
Q

how are blunt ends generated

A

shearing DNA or by cutting with blunt end restriction endonucleases (eg. HindII or SmaI)

35
Q

what does deoxynucleotide terminal transferase do

A

add short poly-T sequences at the 3’-ends of an opened vector.
Add short poly-A sequences to the 3’-ends of DNA to be cloned

36
Q

what is used to seal the blunt ends

A

ligase

37
Q

how is a sticky end formed

A

DNA strands not cut at the centre of the restriction site
staggered cutting leaves overhanging segment of 4 base pairs
5’ G| AA|TTC 3’
3’ CTTAA| G 5’

38
Q

what do sticky ends form

A

transient double helical structures

39
Q

what is used to join sticky ends

A

ligase

40
Q

what happens if different DNA molecules are mixed

A

some ligation will generate new combinations

41
Q

which was the first enzyme discovered

A

Hindll

42
Q

which enzyme can create sticky or blunt ends

A

endonucleases

43
Q

what does a restriction map show

A

positions of restriction sites in sequence of DNA

Applied Genetics (20 decks)

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