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BIOL 2020 > Gene Expression > Flashcards

Gene Expression Flashcards

1
Q

What are constitutive genes?

A

genes that are expressed all the time

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2
Q

What are inducible genes?

A

genes that are turned on at specific times or after certain cues

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3
Q

Most of the regulation occurs at what step?

A

initiation of transcription (happens at a lot of steps)

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4
Q

How is the heat shock gene in E. coli regulated?

A

have variations in core promoter sequence, they recruit a different sigma

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5
Q

What does the normal E. coli promoter vs the heat shock promoter bind?

A

normal binds sigma70, heat shock binds sigma32

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6
Q

How does negative regulation happen?

A

through a repressor, binds and restricts access of promoter to rna polymerase

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7
Q

How does positive regulation happen?

A

through an activator, helps recruit rna polymerase

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8
Q

What enzyme is required for the breakdown of lactose?

A

beta-galactosidase

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9
Q

What do operons typically consist of?

A

gene of interest, promoter (RNA pol binding site), enhancers, operators

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10
Q

What do enhancers do?

A

binding site for transcriptional activators

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11
Q

What do operators do?

A

binding site for transcriptional repressors

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12
Q

What is +1?

A

regulatory sites are in close proxmitity to start site of transcription are called this

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13
Q

What does the lac repressor do in the absence of lactose?

A

forms a homotetramer and binds to either the O3 and O1 sites or the O2 and O1 sites (these are operators), stops rna polymerase from binding

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14
Q

What is the inducer of the lac operon?

A

allolactose, IPTG is the synthetic inducer

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15
Q

How is allolactose made? How does it do its job?

A

lactose is converted into allolactose by LacZ at low levels, binds Lac repressor (Lacl) to make it let go of operator

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16
Q

How is the lac operon regulated through positive regulation?

A

the cAMP receptor protein binds near the promoter when it is bound to cAMP, recruits rna polymerase

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17
Q

When and what makes cAMP?

A

made by adenylate cyclase when glucose levels are low

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18
Q

What are riboswitches?

A

metabolite sensing in the 5-untranslated regions on mRNAs, change shape which influences gene expression

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19
Q

Genes containing riboswitches typically code for what?

A

proteins that are involved in the syntehsis of molecules that are expensive to produce, TPP, FMN etc

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20
Q

What are riboswitches composed of?

A

aptamer (binds metabolite) and expression platform (expression regulator)

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21
Q

How has TPP been shown to alter mRNA structure to inhibit its expression?

A
  1. creates transcriptional terminator
  2. changes structure so ribosome binding site it blocked
  3. alters availibility of splicing zones for mRNA will be non functional
22
Q

What is alternative splicing?

A

joining of different combinations of exons to form cell-specific proteins

23
Q

What does alternative splicing allow?

A

1 gene to make multiple different proteins

24
Q

What is alternative polyadenylation signals?

A

two different cleavage and polyadenylation sites can be used to make different messages and different proteins

25
Q

What is rna interference?

A

cellular mechanism that uses the gene’s own DNA sequence to turn it off

26
Q

What is the first step of rna interference?

A

rna pol ii makes noncoding pri -miRNA that are processed by Drosha microprocessor complex into pre-miRNA

27
Q

In rna interference what happens after pre-miRNA is made?

A

exported to cytoplasm by exportin 5 and processed into double stranded rnas (dsRNAs) by Dicer

28
Q

What happens after dsRNAs are made in RNAi?

A

loaded into RNA induced silencing complex (RISC), one strand is kept (guide) and other is degraded (passenger)

29
Q

After the passenger strand is degraded waht happens next in RNAi?

A

the guide strand is used by RISC to bind mRNA’s by complementarity, if its complenetary than mRNA is cleaved and degraded, if partially complementary then translation repression

30
Q

What are plasmids?

A

naturally occurring,
extra-chromosomal pieces of DNA
that contain all the necessary DNA
sequences to replicate
autonomously in a bacterial host

31
Q

Where is the DNA of interest inserted into plasmid?

A

restriction site

32
Q

What does the insert vs the vector mean?

A

the dna that we inserted, the plasmid

33
Q

What does every plasmid need?

A

origin of replication, antibiotic resistance marker, restriction enzyme sites for the specific cleavage of the plasmid dna to put into the insert

34
Q

What does the antibacterial marker do?

A

allow bacterial cells that contain the plasmid be selected from cells that do not

35
Q

What do they believe restriction endonucleases evolved as?

A

defense mechanism against foreign dna

36
Q

What are the two ways a restriction enzyme can cut? What is the result?

A

staggered, sticky end

same place, blunt end

37
Q

What is the likelihood that a 6 nucleotide long restriction enzyme site is found within a random DNA sequence?

A

1 in 4096 (4^6)

38
Q

Most plasmids used for cloning are engineered to contain what?

A

a polylinker that contains several recognition sequences for commonly used enzymes

39
Q

What can expression vectors do?

A

used to express whatever gene of interest you insert into the polylinker

40
Q

What do expression vectors contain?

A

bacterial promoter and ribosome binding elements from whatever DNA is inserted into the plasmid

41
Q

What is the first step of PCR?

A

strand is heated to 95 degrees to seperate the strands

42
Q

What comes after the denature step in PCR?

A

oligonucleotide primers are added, cooled so they can anneal (50 degree), add Taq DNA polymerase

43
Q

Why Taw dna polymerase in PCR?

A

can withstand higher temperatures without denaturing

44
Q

What is cDNA?

A

dna that is complementary to the mature mRNA (has introns removed)

45
Q

How are cDNAs generated?

A

reverse transcriptase, converts RNA to DNA

46
Q

Once one cDNA is made how can we get more? What is this called?

A

PCR steps, RT-PCR

47
Q

How does sanger sequencing work?

A

primer is annealed to a known sequence upstream of the unkown sequence, then polyermase will add nucleotides until it reaches a chain terminator, can then use electrophoresis to read the segment

48
Q

What are chain terminators?

A

dideoxy nucleotides, has no free OH so nothing can be added after it

49
Q

What is illumina sequencing?

A

similar to sanger sequencing but instead of ddNTPs we use dNTPs that are fluorescent different colours

50
Q

If growing E. coli in a high lactose and high glucose environment what do you expect to find at promoter?

A

No lax repressor bound to operator and no cAMP bound to activatir

BIOL 2020 (8 decks)

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