dna replication, expression, and regulation genetics part 1

What is DNA

Deoxyribonucleic acid - how cells store genetic information, providing instructions for cell production and function

What did Thomas Hunt Morgan's fly experiments show

That chromosomes contained the hereditary material

What is transformation in bacteria

When bacteria can take up genes from their environment

What were the two strains in Griffith's experiment

S (smooth) deadly strain and R (rough) harmless strain of Streptococcus pneumoniae

What did Griffith's experiment show

When heat-killed S cells were mixed with alive R cells and injected into mice, the mice died - showing R cells picked up pathogenic DNA from dead S cells

What did Avery's experiment add to Griffith's work

Added enzymes that destroyed either proteins or DNA - mice with DNA-destroyed batch lived while protein-destroyed batch died, proving DNA contained genes

What is transduction

When a bacteriophage (virus for bacteria) injects its genetic material into a bacterium

What radioisotopes were used in Hershey and Chase experiment

P32 to tag DNA and S35 to tag protein

How to remember Hershey-Chase isotopes

P is 16th letter so 16 × 2 = 32

S is 3 letters after P so 32 + 3 = 35

What did Hershey and Chase experiment show

Radioactive P32 entered bacteria but S35 did not, showing DNA was the genetic material of bacteriophages

What is Chargaff's first rule

The base composition of DNA varied between species

What is Chargaff's second rule

A and T always made up the same percent of bases

C and G always made up the same percent of bases

Who discovered DNA's structure

James Watson (American) and Francis Crick (British)

Whose work did Watson and Crick use

Rosalind Franklin's X-ray crystallography images from Maurice Wilkins' lab

What technique did Rosalind Franklin use

X-ray crystallography to image DNA structure

What are nucleotides composed of

A phosphate, 5-carbon sugar, and a nitrogenous base

What is the 5' end

The end where the phosphate group is bound to the 5th carbon

What is the 3' end

The end where an OH (hydroxyl) group is bound to the 3rd carbon

What is ribose

A 5-carbon sugar with an OH group on the 2nd carbon

What is deoxyribose

A 5-carbon sugar where the OH on the 2nd carbon is removed (de-oxy)

What are phosphodiester bonds

Bonds formed when an O from the 5' phosphate and 3' OH combine, releasing water and linking phosphate to sugar in the sugar-phosphate backbone

What does antiparallel mean

Complementary DNA strands go in opposite directions - if one is 5' to 3', the other is 3' to 5'

What is rise in DNA

The distance between two adjacent bases, which is 0.34 nm

What is pitch in DNA

The distance of a full turn of the helix (roughly every 10 nucleotides), which is 3.4 nm (0.34 × 10)

What are pyrimidines

Nitrogenous bases with a single ring - include C (cytosine) and T (thymine)

RNA has U (uracil) instead of T

What are purines

Nitrogenous bases with 2 rings - include G (guanine) and A (adenine)

Why must a purine bond with a pyrimidine

To maintain constant DNA width and minimize strain

How many hydrogen bonds between A and T

2 hydrogen bonds

How many hydrogen bonds between G and C

3 hydrogen bonds

DNA REPLICATION

What is DNA replication

The process of duplicating DNA so cells can preserve genetic material when dividing

What is the conservative model of replication

One copy consists entirely of original material while the other consists entirely of new material

What is the semiconservative model of replication

Both copies contain one original strand and one new strand - proven correct by Meselson and Stahl

What is the dispersive model of replication

All strands have original and new pieces of DNA mixed together

What isotope was used in Meselson-Stahl experiment

N-15 (heavy nitrogen isotope) in growth medium

What technique was used to analyze DNA in Meselson-Stahl

Equilibrium sedimentation - heavier molecules settle lower on the column

What did Meselson-Stahl find after first generation

One medium band, indicating either semiconservative or dispersive replication

What did Meselson-Stahl find after second generation

Results matched semiconservative model with two bands (one medium, one light)

What does DNA Polymerase III do

Synthesizes DNA from 5' to 3'

Why do polymerases go from 5' to 3'

They use dNTPs and energy comes from breaking phosphate bonds, releasing pyrophosphate, then remaining phosphate binds to 3' end

What happens if ddNTP is used

Dideoxyribose means 3' OH is also removed, so phosphate has nowhere to be added and polymerase stops

What does primase do

Synthesizes RNA primer

What does DNA Polymerase I do

Replaces RNA primers with DNA

How many DNA Polymerases do bacteria have

3 (DNA Polymerase I, II, and III)

How many DNA Polymerases do eukaryotes have

5 (α, β, γ, ϵ, δ)

What does helicase do

Unwinds the DNA

What does topoisomerase do

Relieves stress from supercoiling by unwinding DNA

What do single strand binding proteins do

Prevent separated strands from going back together

What does ligase do

Joins together adjacent segments of DNA

What is the leading strand

The strand with the 5' end closest to helicase where polymerase can replicate continuously toward the helicase

What is the lagging strand

The strand requiring discontinuous replication away from the helicase

Why is the lagging strand more complex

DNA Polymerase needs a preexisting 3' OH to start elongating, but discontinuous replication doesn't provide this

What are Okazaki fragments

Each discontinuous strand on the lagging strand

How do cells recognize newly synthesized vs parent strand in eukaryotes

By the nicks in DNA not yet sealed by ligase

What are mutagens

Substances that damage DNA - include UV radiation, superoxide radicals, and benzene

Why is DNA repair more important than transcription/translation repair

DNA damage is passed on to future cells, but incorrect mRNA and proteins don't ultimately affect cell fate

DNA REPAIR MECHANISMS

What is polymerase exonuclease repair

DNA Polymerase detects wrong base, uses 3' to 5' exonuclease activity to remove it and add correct one

Why does polymerase need to synthesize 5' to 3'

For exonuclease activity - if reversed, after cleaving incorrect base there would be no phosphate bonds to break for energy

What is base excision repair (BER)

Used for non-bulky lesions (DNA damage that doesn't affect helix structure), like chemical modification of one nucleotide

How does BER work

Glycosylases cleave out the base, forming an AP (apurinic/apyrimidinic) site, then repaired by polymerases

What is nucleotide excision repair (NER)

Repairs bulky lesions that alter helix structure, like thymine dimers caused by UV radiation

How does NER differ from BER

NER excises entire nucleotide or segment of nucleotides and is generally more complex

What is direct reversal

An evolutionarily rudimentary version of NER where enzymes can only be used once then must be degraded

When is direct reversal important

In repair after exposure to sunlight

What is non-homologous end joining (NHEJ)

Repairs double strand breaks by annealing the two ends back together - very inaccurate but not costly

What is the downside of NHEJ

Some nucleotides may be lost, leading to potentially harmful deletion

What is homologous end joining (HEJ)

Repairs double strand breaks only after DNA replicates, using homologous strand as template to form Holliday Junction

What do BRCA1/2 genes do

Tumor suppressor genes that play a role in homologous end joining - commonly mutated in breast cancer

TRANSCRIPTION

What is transcription

Converting DNA to RNA using RNA polymerase

What enzyme catalyzes transcription in eukaryotes

RNA polymerase II

What is the promoter

The sequence of DNA where RNA Polymerase binds

What is the TATA box

A sequence just before the promoter where transcription factors bind (Pribnow box in prokaryotes)

What is template DNA

The DNA strand complementary to mRNA that is used for synthesis

What is coding DNA

The DNA strand that is not used for synthesis (doesn't need to debug C++ code)

What are the two methods of termination in prokaryotes

Rho-dependent and Rho-independent

What is attenuation in prokaryotic termination

Formation of a hairpin-like structure by mRNA leading to stalling and dissociation from RNA Polymerase

How does termination work in eukaryotes

RNA Polymerase transcribes a certain sequence, recruits proteins that cleave mRNA off, but keeps transcribing so proteins must chase it down

POST-TRANSCRIPTIONAL MODIFICATION

Do prokaryotic mRNAs undergo modification

Typically no - some ribosomes begin translating before transcription is done

What are introns

Long segments of noncoding DNA in between exons

What are exons

Segments of DNA that code for genes

What is the spliceosome

A RNA-protein complex that pinches out introns and ligates exons

What is splicing

The process of removing introns and joining exons

What is the 5' cap

A modified guanine cap that protects mRNA against degradation

What is the 3' poly-A tail

A string of adenines that protects mRNA against degradation

What makes mRNA "mature"

When it has undergone splicing, 5' capping, and 3' poly-A tail addition

How much of the genome codes for protein

Only three percent

What are some functions of non-coding RNA

In spliceosome (snRNA), regulate gene expression (miRNA, siRNA, lncRNA), repress transposons (piRNA)

TRANSLATION

What is the Shine Dalgarno sequence

A sequence in prokaryotes that makes ribosome binding to mRNA more favorable

What is the Kozak sequence

A sequence in eukaryotes that makes ribosome binding to mRNA more favorable

What is the initiator tRNA

Methionine (AUG) in eukaryotes, formyl-methionine in prokaryotes

What are the three ribosome sites

E (entry), P (peptide/transfer), and A (exit)

What happens at the E site

tRNAs arrive and bind to complementary mRNA codon

What happens at the P site

tRNA transfers its amino acid to the growing polypeptide

What happens at the A site

tRNA exits after transferring amino acid

What are the stop codons

UGA, UAA, UAG

What happens when a stop codon is read

Termination factors arrive at E site and separate ribosome from mRNA

What is a signal sequence/signal peptide

A sequence at the N-terminus (5' end) of some mRNAs

What does the signal sequence bind to

SRP (signal receptor protein) on the ER