tuesday week 3

Question 1

constitutive genes

Answer 1

ALWAYS expressed

Question 2

what are genes called that are always expressed

Answer 2

constitutive

Question 3

regulated genes

Answer 3

genes only expressed in the cell types and conditions in which they are required

Question 4

what are genes called that are not always expressed?

Answer 4

regulated gene

Question 5

describe in situ hybridization

Answer 5

single stranded labeled DNA or RNA probes detect complimentary sequences (of RNA) within tissue, cells, or chromosomes

Question 6

describe/explain what a northern blot is

Answer 6

using RNA not DNA or proteins.

1. seperate strands by mass in agar using electrophoresis
2. transfer RNA strands to nylon membraine
3. add probes (DNA or RNA) which will bind to RNA strands using complementary base pairing
4. probes visualized using x-ray film (dots/blobs we read)

Question 7

cDNA

Answer 7

complementary DNA. DNA synthesized using RNA as a template

Question 8

whatis reverse transcriptase PCR (RT-PCR)

Answer 8

a method of detecting mRNA transcripts using cDNA intermediate

Question 9

steps of reverse transcriptase PCR

Answer 9

1. make cDNA using poly-T prime and reverse transcriptase
2. PCR for mRNA using gene-of-interest specific primers
3. gel electrophoresis to detect gene of interest

Question 10

operon

Answer 10

several genes expressed from a common promoter with a common regulatory DNA sequence (operator)

ONLY in bacteria

Question 11

operator

Answer 11

common regulatory sequence in operon translation

Question 12

polycistronic

Answer 12

a single transcript containing more than one coding sequence that is translated into more than one polypeptide/protein (many rhibosomes translating on the same strand of mRNA)

ONLY in bacteria

Question 13

trp

Answer 13

triptophan

Question 14

trp operon

Answer 14

trp operon is expressed when tryptophan levels are low and repressed when trp levels are high (negative rienforcement loop)

Question 15

low trp

Answer 15

the repressor is not bound by trp and does not bind to DNA. RNA polymerase transcribes the trp operon genes

Question 16

high trp

Answer 16

trp serves as a corepressor and binds to a repressor which has a high afinity for the operator sequence in this state. tryptophan is in repressor which causes conformational change and blocks RNA polymerase from transcribing

Question 17

promoters

Answer 17

general transcription factors + RNA polymerase bind to promoter, this initiates transcription.

promoter is always adjacent and upstream of the coding sequence

promoter activity is directional; and inverted promoter will not function

Question 18

enhancers + silencers

Answer 18

transcription factors (activators or repressors) bind to enhancers and silences and influence the rate of transcription

Question 19

how to enhancers + silencers regulate gene expression

Answer 19

enhancers + silencers regulate transcription through the binding of activators and repressors to their respective enhancer or silencer

enhancers + silencers can regulate at a distance, upstream/downstream, within an intron, even on another chromosome.

enhancer + silencer activity is orientation independent (can face/go any way)

Question 20

mediator protein

Answer 20

connects (physically) enhancers + silencers to protein complex + RNA polymerase

a bridge between transcriptional activators/repressors and RNA polymerase II

Question 21

how the body can modify gene expression during: DNA packing

Answer 21

1. histone tail modifications
2. histone modifying enzymes
3. chromatin remodeling complex

Question 22

histone tail modifications

Answer 22

histone tail modifications: tail modifications can alter afinity of nucleosome to the chromatin fiber or can be bound by regulatory proteins.

• modified using phosphorilation, methylation, etc
• effects tightness/loosness of DNA around nucleosome (histone octomer)

Question 23

histone modifying enzymes

Answer 23

• histone acytl transferase (HATs) increase transcription
• histone deacetylaces (HDACs) reduce transcription

Question 24

chromatin remodeling complex

Answer 24

ATP dependent restructing of DNA relitive to histones (complexes loosen + tighten and/or shift DNA position around nucleosomes (longer shorter ligand DNA))

Question 25

ways gene expression effected by regulation during transcription

Answer 25

1. transcription of a single gene can be regulated by many proteins - lots of things brought together + binded together to regulate a gene 2. a single protein can regulate many genes (sort of like operon in bacteria) - set of different (seperate) genes share a regulatory sequence so one environmental factor (ex. increase in cortosol) will turn on all of these genes at once

Question 26

epigenetic inheritance

Answer 26

the transmission of gene expression from parent to daughter cell(s) WITHOUT altering the DNA sequence

Question 27

how does epigenetic inheritance play into gene expression/why is it important to know about?

Answer 27

if a TRANSCRIPTION FACTOR is part of a positive feedback loop (it activates itself), it can establish 'memory' of cell identity and pass that on without modifying DNA

Question 28

gene expression/regulation during mRNA processing

Answer 28

happens specifically during splicing (usualy). alternative splicing: different combinations of exons are spliced together to produce alternate mRNA's that will produce different protein sequences through splicing of exon(s) or keeping of introns

Question 29

alternative splicing

Answer 29

method of gene regulation. different combinations of exons are spliced together to produce alternate mRNA's that will produce different protein sequences through splicing of exon(s) or keeping of introns

Question 30

how can gene expression be regulated during mRNA stability/degradation steps of transcription

Answer 30

sequences within 3' UTR control the rate of mRNA degradation. the longer the AAAA sequence (tail) the longer it takes to degrade (more stable)

Question 31

how can gene expression be regulated during translation

Answer 31

translation repressor proteins can bind to prevent translation (on mRNA post transcription ofc)

Question 32

micro RNAs (miRNAs)

Answer 32

control mRNA stability and translation

Question 33

RISC

Answer 33

RNA induced silencing complex a single miRNA can inhibit translation of many mRNAs

Question 34

how can gene expression be regulated during protein modification and degradation

Answer 34

proteasome is a protease (enzyme that degrades proteins) (26S) ubiquitin (76aa protein) attaches to side chains of a target protein using ubiquitin ligases

Question 35

what does 26s degrade

Answer 35

- misfolded proteins - proteins that are regulated - unneeded proteins - cleaved protein fragments

Question 36

ubiquitin

Answer 36

76aa protein that attaches to lysine side chains of a target protein using ubiquitin ligases ubiquiting can be thought of as a way cells 'tag' a certain protein for recycling poly-ubiquitinated proteins are unfolded and cleaved by the proteasome