biology 2100 final exam. aka, the most locked in ive ever been. aka, when i lost my sanity.

northern, southern, western blot

used to detect RNA, DNA, protein product

chimera

genetic mix, cells from 2 diff genetic origins

transgenic

foreign DNA inserted in

powerful selection strategies for knockout

homologous recombination, cre-lox, NHEJ

difference between KI and conditional KO

KI is inserting genetic info (replacing?) and conditional KO is a KO that is dependent on location or time

bacteriophage or phage

virus that attacks bacteria

neg/pos control

control resulting in stop of transcription or enhanced transcription

enhancer

regulatory sites in DNA involved in positive control

inducer

small molecules activating gene expression - ex allolactose

repressor protein

transcription factors that help exert negative control

activator protein

transcription factors that help exert positive control

endonuclease

cuts phosphodiester bonds in a polynucleotide chain like RNA or DNA

adaptive immunity

used in nature form of CRISPR/Cas9

restriction enzymes

act like molecular scissors

promoter proximal element

like an enhancer, increases transcription and is close to the promoter

reversible

DNA sequence does not change

x-inactivation

when a gene is silenced in germ cells from a parent to prevent overexpression of a gene - this is not random

genomic imprinting

basically like silencing a gene? - genes are expressed differently depending on if they are inherited by a mother or father

why do we regulate gene expression

to be the most energy efficient we possibly can, specialization, environmental adaptation, make sure cells produce right proteins at right time, etc

coding region

region that contains instructions codes for proteins in transcription

operator function

bind proteins in operons like repressors

negative control hypothesis

something serves as a brake but requires genes for this mechanism

partially diploid

a wild type gene is added to correct a mutant (typically constitutive mutant) in bacteria, only a gene added not whole chromosome and works on trans-acting elements

catabolic pathways

they break things down - lac operon

anabolic pathways

build things up - trp operon

constitutive mutant

always expressing

recombinant DNA

artificially created DNA by combining genetic material from different sources, using tools like restriction enzymes to cut and DNA ligase to join specific DNA pieces

polycistronic mRNA

multiple genes per transcript

allolactose

inducer example

lac operon

inducible operon example

prokaryotes dont have

chromatin or histones or nucleosomes

in trp operon when tryptophan is present …

repressor binds operator bc tryptophan bound repressor changing its confirmation and RNA polymerase was blocked from synthesizing —> no transcription

in trp operon without tryptophan

repressor dissociates from operator and RNA can synthesize

negative

trp regulation

tryptophan is a

corepressor

attenuation

regulatory mechanism that causes premature transcription termination when trp levels are high

trp pathways

anabolic - encodes enzymes for build up

trp default

on

trp change

repressible

CAP (catabolite activator protein)

proteins binding promoter sequences and acting as positive regulators to turn on and activate genes

in CAP cAMP when glucose levels are low

cAMP accumulates because it is produced by adenylyl cyclase which is typically inhibited by glucose, cAMP binds CAP which binds promoter of operon (upstream of RNA pol) and stabilizes RNA binding to increase transcription

in CAP cAMP when glucose levels are high

no cAMP accumulation so CAP doesnt bind promoter, transcription happens at lower rate

CAP cAMP regulation type

positive

lac operon change

inducible

lac operon pathway

catabolic, encodes for breaking down something

lac operon default

off

lac operon

inducible operons have proteins that bind to activate transcription depending on local environment and cell needs

lacZ

codes for B-galactosidase

B-galactosidase

breaks lactose into glucose and galactose for glycolysis

LacY

codes for permease

permease

brings lactose into cell

lacI

codes for repressor

conditions for lac operon

lactose present, glucose not

part of lac operon that repressor and lactose make

negatively inducible

part of lac operon that CAP enhances

positive regulation/activation

yes glucose no lactose

transcription blocked

yes glucose yes lactose

transcription at low rate

no glucose yes lactose

transcription at high rate

no glucose no lactose

no transcription

arabinose operon - normal

catabolizes arabinose and involves regulatory protein araC bound for gene to catabolize

arabinose operon no arabinose

regulatory protein araC acts as inhibitor for RNA polymerase

ara operon w/ arabinose

arabinose binds araC and alters its conformation so that it can stimulate RNA pol (stabilize i assume)

araC control

positive and negative because it inhibits RNA pol without arabinose and stimulates RNA pol with arabinose

how to replica plate

grow master plate of mutated e coli colonies on medium containing glucose as energy source - then press velvet block and transfer colonies from cell onto lactose plate and observe

3 mutant types

LacZ-, lacY-, constitutive

lacZ-

no B-galactosidase, no glucose and galactose, no glycolysis - color on plate

lacY-

no permease, no lactose in cell, die on lactose plate

constitutive

expression without regulation, express permease and B-galactosidase even without lactose - WASTE!

partially diploid cannot help

operator constitutive mutants - they are cis acting

partially diploid can help

lacI constitutive mutants - trans acting

similarity P and E

promoter upstream/before 5’ end of newly synthesizng RNA

similarity P and E

promoter is where RNA pol binds DNA and begins transcription

similarity P and E

TFs can enhance or repress transcription by binding regulatory sequences

operator

regulatory sequence

similarity P and E

have fitness advantages

similarity P and E

negative and positive control and can regulate transcription

similarity P and E

have DNA regulatory sequences

similarity P and E

can change protein activity by adding or removing a phosphate group

only P

lacks nucleus

only P

DNA in cytoplasm

only P

simultaneous transcription and translation

only P

primarily regulates gene expression at transcriptional level

only P

can have multiple stop and start codons in same mRNA, contain sequences that can make diff proteins

only P

regulatory sequences are promoter proximal

only P

has operons

only E

has nucleus

only E

DNA confined to nucleus

only E

transcription in nucleus THEN translation in cytoplasm

only E

has many levels of gene regulation

only E

coding sequences in introns and exons

only E

exons expressed and code for amino acids

only E

introns between exons but removed before translation

only E

regulatory sequences can be close or far

only E

some introns have regulatory sequences that serve as binding sites or TFs

only E

chromatin

chromatin

DNA and histones

chromosomes

made of chromatin

histones have

many arginines and lysines

charge of arginines and lysines and histones

positive

charge of DNA sugar phosphate backbone (due to phosphate groups)

negative