cell bio 2

How are different traits inherited from one generation to the other?

• Traits are passed through genes, which are segments of DNA.

• Offspring inherit genes/alleles from their parents

What was the experimental setup for Frederick Griffith’s experiment?

• Used S strain bacteria: smooth, capsule, deadly to mice.

• Used R strain bacteria: rough, no capsule, harmless.

• Heat-killed S bacteria + live R bacteria → mouse died and live S bacteria were found.

What was the major conclusion from Griffith’s experiment?

• A substance from dead S bacteria transformed R bacteria into S bacteria.

• This was called the transforming principle.

What was the experimental setup for Avery, MacLeod, and McCarthy’s experiment?

• Isolated DNA, RNA, and proteins from S bacteria.

• Destroyed each type separately using enzymes.

• Tested whether R bacteria could still be transformed.

What did Avery, MacLeod, and McCarthy’s experiment conclude?

• Destroying DNA stopped transformation.

• Therefore, DNA was the transforming principle.

What is the chemical substance that carries traits?

DNA (deoxyribonucleic acid)

What is DNA made up of?

• DNA is a polymer made of nucleotides.

• It contains sugar, phosphate, and nitrogenous bases.

What are nucleotides? What bond connects them?

• Nucleotides are the building blocks of DNA and RNA.

• Nucleotides are connected by phosphodiester bonds.

what are components of a nucleotide?

• 5-carbon sugar

• Phosphate group

• Nitrogenous base

What is special about the 5′ and 3′ positions?

• The 5′ carbon is associated with the phosphate.

• The 3′ carbon has an OH group where the next nucleotide is added.

• DNA is synthesized 5′ → 3′.

What are the two types of bases in DNA?

• Purines: A and G

• Pyrimidines: C and T

What is Chargaff’s rule?

• A = T

• G = C

• A always pairs with T; G always pairs with C.

What was concluded from Rosalind Franklin’s diffraction pattern?

• DNA has a helical structure.

• It provided evidence about DNA’s dimensions and arrangement of phosphate groups.

What was Watson and Crick’s DNA model?

• DNA is a double helix.

• Two strands run antiparallel.

• Bases are inside; sugar-phosphate backbones are outside.

What resides in the inner core (inside) of DNA? What makes the backbone (outside)?

• Inside: nitrogenous bases.

• Outside: alternating sugar and phosphate groups.

What are the features of base pairing in DNA?

• A pairs with T using 2 hydrogen bonds.

• G pairs with C using 3 hydrogen bonds.

• One purine pairs with one pyrimidine.

• The strands are complementary and antiparallel.

What experiment showed DNA replicated semi-conservatively?

• Meselson and Stahl experiment.

• Used heavy ¹⁵N and normal ¹⁴N.

• DNA density after replication showed that each new DNA molecule contained an old and new strand.

What does semi-conservative mean?

• Each new DNA molecule contains:

  • 1 original strand

  • 1 newly made strand

DNA replication is catalyzed by what enzyme?

DNA polymerase

What is the template strand?

The existing DNA strand used as a guide to make the new complementary strand

What is a growing strand?

The new DNA strand being synthesized by DNA polymerase.

What enzyme unwinds DNA?

• Helicase

• Breaks hydrogen bonds and separates the DNA strands.

What are the key features of DNA polymerase?

• enzymes that builds new DNA strands by matching nucleotides to an existing template strand

• Adds nucleotides 5′ → 3′.

• Has high fidelity (accuracy).

• Can proofread and remove many incorrect nucleotides.

• Requires a primer to begin synthesis.

What is the difference between nucleotides and nucleosides?

• Nucleoside = sugar + base

• Nucleotide = sugar + base + phosphate

Which bases are purines and pyrimidines?

• Purines: Adenine (A), Guanine (G)

• Pyrimidines: Cytosine (C), Thymine (T)

What are phosphodiester bonds?

• Covalent bonds connecting nucleotides together.

• Form the sugar-phosphate backbone.

What does bidirectional DNA replication mean?

• Replication proceeds in two directions from the origin.

• Two replication forks move away from the origin.

What are an origin, replication fork, and replicon?

• Origin: location where DNA replication begins.

• Replication fork: Y-shaped area where DNA is being copied.

• Replicon: section of DNA replicated from one origin.

What are the key molecules/enzymes in DNA replication?

• Helicase: unwinds DNA.

• SSB proteins: keep DNA strands separated.

• Topoisomerase: reduces twisting/tension.

• Primase: makes RNA primers.

• DNA polymerase: adds DNA nucleotides.

• DNA ligase: joins DNA fragments.

• Sliding clamp: keeps DNA polymerase attached to DNA.

What are the leading and lagging strands?

• Leading: made continuously.

• Lagging: made in short pieces because DNA synthesis only occurs 5′ → 3′.

What are Okazaki fragments? Why are they created?

• Short DNA pieces made on the lagging strand.

• Created because DNA polymerase can only synthesize 5′ → 3′.

• Ligase joins them together.

What mechanism prevents DNA shortening at the end of the lagging strand?

• Telomerase helps maintain chromosome ends.

• It extends telomeres so important DNA is not progressively lost.

What are telomeres and telomerase?

• Telomeres: repetitive DNA at chromosome ends that protect them.

• Telomerase: enzyme that extends telomeres.

What are common DNA lesions?

• Thymine dimers from UV radiation.

• Depurination: loss of a base.

• Deamination: chemical alteration of a base.

• Mismatch: incorrect base pairing.

• Single- or double-strand breaks.

What are four DNA repair mechanisms?

• Direct repair: directly reverses certain damage.

• Base excision repair: removes damaged individual bases.

• Nucleotide excision repair: removes a larger damaged section.

• Mismatch repair: fixes incorrect base pairs left after replication.

How did Beadle and Tatum develop the one gene–one enzyme hypothesis?

• Mutated Neurospora using radiation.

• Some mutants could not make specific substances.

• They connected specific genes with specific enzymes.

• Led to the one gene–one enzyme hypothesis.

What does each gene make?

• A gene produces a functional product.

• This can be a protein or functional RNA.

What is the central dogma?

• DNA → RNA → Protein

• DNA → RNA = transcription

• RNA → protein = translation

What is making RNA from DNA called?

Transcription

What is making protein from RNA called?

Translation

Where is RNA made? Where is protein made?

• Eukaryotes: RNA is mainly made in the nucleus; protein is made by ribosomes.

• Prokaryotes: both occur in the cytoplasm.

What is a codon?

• A group of 3 mRNA nucleotides.

• Specifies an amino acid or a stop signal.

What are mRNA and tRNA?

• mRNA: carries genetic instructions to the ribosome.

• tRNA: carries amino acids to the ribosome.

What is the function of the ribosome?

• Reads mRNA.

• Positions tRNAs.

• Builds the protein by forming peptide bonds.

What is the chemical difference between DNA and RNA?

• DNA has deoxyribose; RNA has ribose.

• DNA uses T; RNA uses U.

• DNA is usually double-stranded; RNA is usually single-stranded.

What are some secondary and tertiary structures of RNA?

• Secondary: hairpins, stem-loops, internal loops.

• Tertiary: complex 3D folding of RNA, such as tRNA.

What are the key players in transcription?

• RNA polymerase: makes RNA.

• Promoter: DNA region where transcription begins.

• Transcription factors: help regulate transcription.

• NTPs: building blocks of RNA.

What are the three steps of transcription?

1. Initiation: RNA polymerase binds promoter.

2. Elongation: RNA strand is built.

3. Termination: transcription stops and RNA is released.

What is the directionality of RNA?

• RNA is made 5′ → 3′.

• New nucleotides are added to the 3′ end.

What are post-transcriptional modifications?

• 5′ cap

• Poly-A tail

• RNA splicing

Does RNA processing happen in both eukaryotes and prokaryotes?

• Extensive mRNA processing is mainly characteristic of eukaryotes.

• Prokaryotes can have RNA processing, but generally do not use the same 5′ cap/poly-A/splicing system.

What happens at the 5′ and 3′ ends of mRNA?

• 5′ end: receives a 5′ cap.

• 3′ end: receives a poly-A tail.

• These help protect mRNA and help with translation.

What happens to introns?

• Introns are removed from pre-mRNA.

• Exons are joined together.

What mechanism splices introns?

• The spliceosome removes introns and joins exons.

What are snRNPs?

• Small nuclear ribonucleoproteins.

• Made of snRNA + proteins.

• Help form the spliceosome and remove introns.

What happens to introns finally?

• They are usually released and degraded/recycled by the cell.

How many amino acids are present?

• There are 20 standard amino acids used to make proteins.

How are amino acids made from nucleotides?

• Three nucleotides form a codon.

• Each codon specifies an amino acid.

• The ribosome uses the codons to determine the amino acid sequence.

What is a start codon?

• AUG

• Usually codes for methionine.

• Signals where translation begins.

What is a stop codon?

• UAA, UAG, UGA

• Signals the end of translation.

What are the features of the genetic code?

• Triplet: 3 bases per codon.

• Degenerate: multiple codons can code for the same amino acid.

• Unambiguous: each codon specifies only one amino acid or stop.

• Nearly universal: used by most organisms.

What is the function of tRNA?

Carries a specific amino acid to the ribosome

What is the anticodon domain?

• Three bases on tRNA that pair with a complementary mRNA codon

What is the acceptor stem?

• The 3′ end of tRNA.

• The amino acid attaches to this end

• end labeled ‘A’

What is wobble base pairing?

• Flexible pairing at the third base of a codon.

• Allows one tRNA to recognize multiple codons.

What is charging of tRNA?

• Attaching the correct amino acid to tRNA.

• Done by aminoacyl-tRNA synthetase.

• Requires ATP.

What are the three ribosome sites?

• A site: incoming tRNA.

• P site: holds tRNA carrying growing protein.

• E site: tRNA exits.

What are the steps of translation?

1. Initiation: ribosome finds start codon.

2. Elongation: amino acids are added.

3. Termination: stop codon causes protein release.

What catalyzes peptide bond formation?

• The ribosome's peptidyl transferase center, primarily rRNA.

What structure do many antibiotics bind to in bacteria?

• Many antibiotics target the 70S bacterial ribosome.

• They interfere with protein synthesis, which can stop bacterial growth or kill the bacteria.

What is PCR? What is its purpose?

• Polymerase Chain Reaction.

• Amplifies a specific DNA sequence to make millions of copies.

What components are needed for PCR?

• Template DNA.

• Forward and reverse primers.

• DNA polymerase.

• dNTPs.

• Buffer/Mg²⁺.

• Water.

What are the steps of PCR?

1. Denaturation: DNA strands separate.

2. Annealing: primers attach.

3. Extension: DNA polymerase copies DNA.

What are uses of PCR?

• Disease testing.

• Forensics.

• Genetic testing.

• Research.

• DNA sequencing preparation.

What are restriction enzymes?

• Enzymes that cut DNA at specific sequences.

What is gel electrophoresis?

• Separates DNA fragments by size.

• DNA moves toward the positive electrode.

• Smaller fragments move farther.

• DNA is visualized using DNA-binding dyes.

How do restriction enzymes cut DNA?

• Recognize specific DNA sequences.

• Cut the DNA backbone.

• Can create sticky ends or blunt ends.

What are plasmids?

• Small, usually circular DNA molecules found in bacteria.

• Used as vectors to carry foreign DNA.

What special feature helps select plasmids?

• Often contain an antibiotic-resistance gene.

• Only bacteria with the plasmid survive antibiotic selection.

What are the steps to express recombinant protein?

1. Isolate/amplify gene.

2. Insert gene into plasmid.

3. Put plasmid into host cell.

4. Select cells containing plasmid.

5. Express the gene.

6. Collect/purify protein.

What are uses of recombinant proteins?

• Insulin.

• Medicines.

• Vaccines.

• Industrial enzymes.

• Research and diagnostics.

What technologies can knock down a gene?

• RNA interference (RNAi/siRNA)

• Antisense RNA

• CRISPRi

What is an industrial application of gene knockdown?

• Reduce production of unwanted products.

• Increase useful product production.

• Modify microorganisms for chemicals, medicines, or fuels.

What are biofuels?

• Fuels made from biological materials.

• Examples: ethanol, biodiesel, biogas.

How is CRISPR-Cas9 used to knock down a gene or insert DNA?

• Guide RNA directs Cas9 to a specific DNA sequence.

• Cas9 cuts the DNA.

• Repair can disrupt the gene.

• A DNA template can sometimes be provided to help insert new DNA.

What was the old method of DNA sequencing?

• Sanger sequencing (chain-termination method).

• Uses DNA polymerase and special nucleotides to determine DNA sequence.

What was the use of ddNTPs?

• ddNTPs stop DNA synthesis because they lack a 3′-OH group.

• Different ddNTPs stop DNA at different positions, allowing the sequence to be determined

What is next-generation DNA sequencing?

• High-throughput sequencing that reads millions of DNA fragments at once.

• Much faster for large amounts of DNA than traditional Sanger sequencing.

What is RNA-seq?

• Uses sequencing to study the RNA transcripts in a cell.

• Shows which genes are being expressed and their relative expression levels.

What is a disadvantage of studying transcriptomics?

• RNA levels can change depending on cell type, environment, and time.

• RNA levels also do not always equal the amount or activity of protein.

How many genes does the human genome have?

• About 20,000 protein-coding genes.

What is proteomics?

• Study of the entire set of proteins in a cell or organism.

• Examines protein amounts, structures, functions, and interactions.

What are protein families?

• Groups of proteins with a common evolutionary ancestor.

• Often have similar sequences, structures, or functions.

What comprises the human genome besides coding sequences?

• Regulatory DNA.

• Introns.

• Noncoding RNA genes.

• Repetitive DNA.

• Intergenic regions.

• Telomeres and centromeres.

What does it mean to have homologous proteins?

• Proteins that evolved from a common ancestral protein.

• They may have similar or different functions.

What is metagenomics?

• Study of DNA collected directly from an environmental or biological sample.

• Can identify and study many microorganisms without growing each one separately.

What are SNPs?

• Single Nucleotide Polymorphisms.

• Differences in a single DNA nucleotide between individuals.

How are SNPs used to identify individuals?

• People have different combinations of SNPs.

• Analyzing many SNPs together can create a genetic profile that distinguishes individuals.

How can genomics revolutionize medicine?

• Helps identify disease-associated genetic variants.

• Allows personalized treatment.

• Helps predict medication responses.

• Can improve disease diagnosis and drug development.

How can genomics help better treat cancer?

• Identifies mutations in a person's tumor.

• Helps classify the cancer.

• Can identify targetable mutations.

• Helps doctors choose treatments that target specific molecular changes.

• Can help track treatment resistance and changes in the tumor.