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BMSC 210 > Lecture 13: Biotechnology > Flashcards

Lecture 13: Biotechnology Flashcards

1
Q

genetic engineering

A

in vitro techniques to alter genetic material from prokaryotes

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2
Q

where does biobrick assembly occur?

A

test tube

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3
Q

heterologous expression

A

expressing a gene in a different host

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4
Q

PCR

A

rapid amplification in DNA sequence copies

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5
Q

thermocycler

A

automated PCR machine

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6
Q

Quantitative PCR

A

amplifies RNA after conversion to DNA

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7
Q

PCR steps

A
  1. denaturation
  2. flanking/binding
  3. add DNA polymerase
  4. Heat & cool
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8
Q

reverse transcription PCR

A

makes DNA from mRNA template

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9
Q

what does reverse transcription PCR do?

A

produces intron free eukaryotic gene

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10
Q

what end is the primer added to in PCR?

A

5’

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11
Q

Gel electrophoresis

A

separates DNA based on size & charge

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12
Q

what electrode do nucleic acids migrate to?

A

positive

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13
Q

what size of molecules move faster in gel electrophoresis

A

small molecules

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14
Q

what does ethidium bromide do in gel electrophoresis?

A

is used to visualize the DNA fragment bands

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15
Q

will the same DNA cut with different restriction enzymes have the same or different banding patterns?

A

different

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16
Q

nucleic acid hybridization

A

DNA / RNA from 2 diff sources form a hybrid double helix

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17
Q

nucleic acid probe

A

segment of DNA strand that has a predetermined identity

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18
Q

southern blot

A

DNA is in the gel & probe is RNA/DNA

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19
Q

northern blot

A

RNA is in the gel & the probe is DNA/RNA

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20
Q

synthetic DNA usage

A

used for primers, probes & site directed mutagenesis

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21
Q

what is synthetic DNA made of

A

oligonucleotides of 100 bases

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22
Q

molecular cloning

A

movement of a gene from original source to small/manipulable genetic element

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23
Q

what does molecular cloning result in?

A

recombinant DNA

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24
Q

recombinant DNA

A

molecule containing DNA from different sources

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25
Q

what 4 enzymes are used for cloning?

A
  1. restriction endonucleases
  2. DNA ligase
  3. reverse transcriptase
  4. DNA polymerase
26
Q

what does DNA ligase join together

A

joins 5’ phosphate & 3’ OH of adjacent nucleotides

27
Q

DNA polymerase

A

5’ to 3’ activity

28
Q

what do restriction endonucleases do

A

cuts DNA

29
Q

where are restriction endonucleases more commonly found

A

prokaryotes

30
Q

Type II restriction endonucleases

A

cleaves DNA within sequence
- most useful
- palindrome

31
Q

Type I

A

cuts DNA randomly & not within sequence

32
Q

Type III

A

cleaves outside of sequences

33
Q

recombineering

A

allows foreign DNA to be inserted into vectors/chromosome

34
Q

what was recombineering originally developed in?

A

saccharomyces cerevisiae

35
Q

where do cloning vectors replicate

A

the host

36
Q

why are plasmids used as cloning vectors?

A

small size & they have selectable markers

37
Q

pUC19 uses

A

insertional inactivation

38
Q

insertional inactivation

A

lacZ is inactivated & B-galactosidase is not produced

39
Q

blue vs white colonies

A

blue: vector is not inserted with foreign DNA
white: vector is inserted with foreign DNA

40
Q

PCR cloning vectors

A

plasmids specifically for cloning DNA products made by taq

41
Q

where are shuttle vectors replicated & maintained

A

replicate & be maintained in E. coli & yeast

42
Q

yeast artificial chromosomes (YACs)

A

chromosomes that clone directly into yeast

43
Q

properties of a good host for cloning

A
  1. non-pathogenic
  2. rapid growth
44
Q

electroporation

A

electric pulse is applied to form pores

45
Q

microinjection

A

gene gun inserts into cells

46
Q

expression vectors

A

vectors that control expression of genes

47
Q

why are eukaryotic genes that contain introns easy to isolate?

A

because of poly (A) tails

48
Q

fusion proteins

A

join target & carrier proteins

49
Q

site directed mutagenesis

A

introduces mutations at a precise location

50
Q

what can site directed mutagenesis be used for?

A

assess amino acids activity in a protein

51
Q

where does site directed mutagenesis occur?

A

in vitro

52
Q

how many base DNA oligonucleotides can be obtained through site directed mutagenesis

A

12 - 40

53
Q

cassette mutagenesis

A

synthetic fragments that can change base pair changes

54
Q

gene disruption

A

inserts cassettes in middle of a gene

55
Q

knockout mutation

A

loss of function of a gene in which cassette is inserted

56
Q

reporter gene

A

encodes protein easy to detect & assay

57
Q

example of a reporter gene

A

green fluorescent proteins

58
Q

operon fusions

A

signals are fused with transcriptional signals of another gene

59
Q

protein fusions

A

2 proteins are fused & share same transcriptional & translational start/stop

60
Q

what do protein fusions yield?

A

1 hybrid polypeptide

61
Q

genome editing

A

uses CRIPSR from streptococcus pyogenes to alter eukaryotic genomes

BMSC 210 (16 decks)

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