DNA Replication and Transcription
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Questions and Answers

During DNA replication, DNA polymerase reads the template strand in which direction?

  • It depends on whether it is the leading or lagging strand.
  • 5′ → 3′
  • 3′ → 5′ (correct)
  • Both 5′ → 3′ and 3′ → 5′

If a segment of DNA undergoes two rounds of semiconservative replication starting with only heavy nitrogen (15N), what percentage of the resulting DNA helices would contain at least one strand of the original 15N DNA?

  • 100%
  • 0%
  • 25%
  • 50% (correct)

Mismatch repair (MMR) is crucial for correcting errors during DNA replication. What is the primary function of MMR?

  • Repairing double-strand breaks in DNA.
  • Correcting errors during transcription.
  • Adding telomeres to the ends of chromosomes.
  • Replacing mismatched DNA nucleotides. (correct)

DNA synthesis is best described by which type of chemical reaction?

<p>Condensation, releasing water molecules. (D)</p> Signup and view all the answers

Activators and repressors regulate transcription by affecting the binding of what molecule?

<p>RNA Polymerase (B)</p> Signup and view all the answers

During reverse transcription PCR (RT-PCR), what must be done to the generated cDNA sequence to determine the sequence found in the original mRNA?

<p>Reverse complement the cDNA sequence, replacing T with U. (C)</p> Signup and view all the answers

Alternative splicing allows one gene to code for multiple proteins. What best describes this process?

<p>It allows different combinations of exons to be included in the final mRNA. (D)</p> Signup and view all the answers

What is the most likely effect of increased competition from cap analogs during translation?

<p>Decreased cap-dependent translation (B)</p> Signup and view all the answers

Small nuclear RNAs (snRNAs) play a critical role in which of the following processes?

<p>Splicing pre-mRNA in the nucleus (B)</p> Signup and view all the answers

You are studying mRNA degradation rates. Which method would be most appropriate to quantitatively track mRNA levels over time?

<p>qRT-PCR (A)</p> Signup and view all the answers

Which histone modification is typically associated with increased gene transcription?

<p>Histone Acetylation (D)</p> Signup and view all the answers

In somatic cells, the TERT gene is often not expressed because it's stored in heterochromatin. How does heterochromatin affect gene expression?

<p>Heterochromatin limits access of RNA polymerase to the DNA. (A)</p> Signup and view all the answers

In nonreducing SDS-PAGE, a wild-type protein (WT) exists as a homodimer, while a mutant protein exists as a monomer. How would their migration differ?

<p>The WT would move slower than the mutant. (A)</p> Signup and view all the answers

In gel electrophoresis, if you are using a northern blot to analyze mRNA, how do mRNA fragment size and migration distance relate?

<p>Smaller fragments migrate further down the gel. (A)</p> Signup and view all the answers

Which set of enzymes is required for the construction of vector DNA for cloning?

<p>Reverse transcriptase, DNA polymerase, restriction enzymes, and ligase. (D)</p> Signup and view all the answers

Gene duplication is a key evolutionary mechanism. What is a typical outcome of gene duplication events over long periods?

<p>The duplicated genes mutate and acquire different functions. (D)</p> Signup and view all the answers

In bacterial DNA transfer, what is the key difference between conjugation and transformation?

<p>Conjugation involves direct transfer of DNA between bacteria. (C)</p> Signup and view all the answers

A protein is found to be hyperphosphorylated in diseased cells. What strategies could restore normal protein function?

<p>Increase phosphatase activity or decrease kinase activity. (D)</p> Signup and view all the answers

How do antisense oligonucleotides (ASOs) affect pre-mRNA splicing and protein production to treat diseases like spinal muscular atrophy (SMA)?

<p>They alter pre-mRNA splicing to increase functional protein production. (C)</p> Signup and view all the answers

What is the effect of mutations in enhancer regions on gene transcription and disease risk?

<p>They increase transcription by allowing more transcription activators to bind. (D)</p> Signup and view all the answers

If a cell line exhibits increased cap analog competition, how would this affect cap-dependent and IRES-mediated translation?

<p>Cap-dependent translation would decrease, while IRES-mediated translation would remain unaffected. (D)</p> Signup and view all the answers

A researcher wants to express a eukaryotic gene in bacteria. Why is cDNA preferable to genomic DNA for this purpose?

<p>cDNA lacks introns, so it does not require splicing by the bacterial cell. (A)</p> Signup and view all the answers

Which of the following processes is most directly affected by histone acetylation?

<p>Gene transcription rates (A)</p> Signup and view all the answers

In somatic cells, the TERT gene is typically silenced by packaging in heterochromatin. How does this affect the availability of the TERT gene for transcription?

<p>Heterochromatin prevents RNA polymerase from accessing the TERT gene, leading to transcriptional silencing. (B)</p> Signup and view all the answers

You run a nonreducing SDS-PAGE. Under nonreducing conditions, what accounts for the difference in migration between a wild-type homodimer protein and a mutant monomer protein?

<p>The wild-type homodimer migrates slower due to intact disulfide bonds linking its subunits. (C)</p> Signup and view all the answers

In Northern blotting, smaller mRNA fragments migrate further down the gel. What characteristic of mRNA molecules is being exploited in this separation technique?

<p>Molecular weight (A)</p> Signup and view all the answers

A researcher aims to confirm that a specific gene is actively expressed in a tissue sample. Which method would be most appropriate for detecting the presence of the gene's mRNA transcript?

<p>Hybridization with a labeled RNA probe (A)</p> Signup and view all the answers

Which enzyme is NOT required to construct a vector for cloning a gene of interest?

<p>RNA polymerase (B)</p> Signup and view all the answers

What is the likely effect of increased DNA methylation in a gene promoter region?

<p>Reduced gene transcription due to chromatin condensation. (D)</p> Signup and view all the answers

In Alzheimer's disease, hyperphosphorylation of the protein tau leads to neurofibrillary tangles. Which therapeutic strategy would directly address this issue?

<p>Inhibition of kinases that phosphorylate tau. (D)</p> Signup and view all the answers

What is the crucial difference between DNA polymerase and RNA polymerase in molecular biology?

<p>DNA polymerase replicates DNA; RNA polymerase transcribes DNA into RNA. (B)</p> Signup and view all the answers

During DNA replication, DNA polymerase adds nucleotides in which direction in relation to the template strand?

<p>Reads 3' → 5', synthesizes 5' → 3'. (C)</p> Signup and view all the answers

If a double-stranded DNA molecule undergoes three rounds of semiconservative replication, what proportion of the resulting DNA molecules would contain at least one strand from the original molecule?

<p>1/4 (C)</p> Signup and view all the answers

Double-strand breaks (DSBs) are a critical type of DNA damage. Where do these breaks primarily occur, and what enzymatic activity is required for their repair?

<p>In the deoxyribose-phosphate backbone; requires phosphodiester bond formation. (B)</p> Signup and view all the answers

Mismatch repair primarily targets errors in which type of molecule and what is its main function?

<p>DNA; replaces mismatched DNA nucleotides to correct replication errors. (A)</p> Signup and view all the answers

What type of chemical reaction is DNA synthesis best described as, and what byproduct is released in the process?

<p>Condensation, releasing water. (A)</p> Signup and view all the answers

Activators and repressors regulate transcription by influencing the binding ability of what molecule?

<p>RNA Polymerase. (A)</p> Signup and view all the answers

In reverse transcription PCR (RT-PCR), how do you determine the original mRNA sequence from the generated cDNA sequence?

<p>The cDNA sequence is the reverse complement of the mRNA sequence, with T replaced by U. (C)</p> Signup and view all the answers

Alternative splicing is a crucial process in gene expression. What does alternative splicing allow a single gene to do?

<p>Produce multiple proteins by combining different exon combinations. (B)</p> Signup and view all the answers

How does increasing the competition from cap analogs affect translation, and why?

<p>It decreases cap-dependent translation by limiting ribosome binding to mRNA. (D)</p> Signup and view all the answers

MicroRNAs (miRNAs) are essential regulators of gene expression. By what mechanism do miRNAs typically function?

<p>Binding to complementary mRNA to block translation or degrade the mRNA. (A)</p> Signup and view all the answers

To quantitatively track mRNA levels over time, which method would be most effective to analyze degradation rates?

<p>qRT-PCR to quantify cDNA levels converted from mRNA. (C)</p> Signup and view all the answers

Which histone modification is generally associated with increased gene transcription activity and how does it affect chromatin structure?

<p>Acetylation, which loosens chromatin. (A)</p> Signup and view all the answers

How does packaging the TERT gene in heterochromatin in somatic cells affect its expression?

<p>Prevents RNA polymerase from accessing the TERT gene, thus silencing its expression. (C)</p> Signup and view all the answers

In nonreducing SDS-PAGE, how would the migration of a wild-type protein existing as a homodimer differ from that of a mutant protein existing as a monomer?

<p>The homodimer would migrate slower due to intact disulfide bonds. (D)</p> Signup and view all the answers

In gel electrophoresis, if you are using a northern blot to analyze mRNA, what determines how far an mRNA fragment migrates?

<p>The size of the mRNA fragment. (B)</p> Signup and view all the answers

What set of enzymes is essential for constructing vector DNA for cloning a gene of interest?

<p>Reverse transcriptase, DNA polymerase, restriction enzymes, and ligase. (D)</p> Signup and view all the answers

Gene duplication is a significant evolutionary event. Over extended periods, what commonly happens to duplicated genes?

<p>They accumulate mutations, potentially leading to new or altered functions. (D)</p> Signup and view all the answers

What is the key distinction between conjugation and transformation in bacterial DNA transfer?

<p>Conjugation involves direct DNA transfer between bacterial cells, while transformation involves the uptake of foreign DNA from the environment. (B)</p> Signup and view all the answers

In Alzheimer's disease, hyperphosphorylation of tau protein leads to neurofibrillary tangles. Which therapeutic strategy would directly address this issue to restore normal protein function?

<p>Increase phosphatase activity to reduce tau phosphorylation. (C)</p> Signup and view all the answers

Flashcards

Replication vs. Transcription

DNA polymerase replicates DNA, while RNA polymerase transcribes DNA into RNA.

DNA Replication Direction

DNA polymerase reads 3' → 5' and synthesizes 5' → 3', linking the 3' OH of the growing strand to the 5' phosphate of incoming nucleotides.

Semiconservative Replication

Each round of replication halves the amount of original (15N) strands.

Mismatch Repair (MMR)

Fixes DNA replication errors by replacing mismatched DNA nucleotides, but does not act on RNA.

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Activators vs. Repressors

Activators increase transcription by helping RNA polymerase bind, while repressors inhibit transcription by blocking RNA polymerase.

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Poly-A Tail & 5' Cap

Poly-A tail (3' end) protects mRNA and aids nuclear export, while the 5' Cap prevents degradation and assists in ribosome binding.

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Genetic Code Properties

Redundant (degenerate), meaning multiple codons code for the same amino acid, but not ambiguous as each tRNA carries only one amino acid.

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Mutations

Silent (no amino acid change), missense (swaps one amino acid), nonsense (creates a premature stop codon), and frameshift (insertion/deletion shifts the reading frame).

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rRNA & Ribosomes

rRNA builds ribosomes and catalyzes peptide bond formation.

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miRNA Gene Silencing

miRNAs bind to complementary mRNA, blocking translation or degrading mRNA.

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Euchromatin vs. Heterochromatin

Euchromatin (acetylated histones) is open and active in transcription, while heterochromatin (deacetylated/methylated histones) is tightly packed and has low transcription.

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Histone Modifications

Acetylation loosens chromatin, increasing transcription, while deacetylation condenses chromatin, decreasing transcription.

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TERT Regulation in Somatic Cells

Somatic cells rarely express TERT, so it's stored in heterochromatin, limiting RNA polymerase access.

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Northern Blotting

mRNA separation by size; smaller fragments move further down the gel.

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DNA Methylation

Occurs on cytosine (C), reducing transcription, not increasing it.

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Telomeres

Telomeres shorten with each division and require telomerase.

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Gene transcription in somatic cells

In somatic cells, genes that are rarely transcribed, like TERT, are typically stored in heterochromatin, a tightly packed DNA form that limits accessibility to RNA polymerase and reduces gene expression.

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Repressor action

When repressors act, they inhibit transcription by preventing RNA polymerase from binding to the promoter.

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Activator action

Conversely, activators facilitate transcription by helping RNA polymerase bind more efficiently.

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Knockout organisms

Knockout organisms have an intentionally inactivated or nonfunctional gene to identify its biological role.

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Cap-Dependent vs. Cap-Independent Translation

Cap-dependent translation decreases with increased cap analog competition. IRES-mediated translation is unaffected by cap analogs.

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snRNA Function

Splices pre-mRNA in the nucleus to produce mature mRNA.

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cDNA Expression

cDNA lacks introns and does not need splicing but still results in the same protein product as the original gene.

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mRNA Half-Life Analysis

Convert mRNA to cDNA and measure degradation over time using quantitative real-time PCR (qRT-PCR).

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Vector DNA Construction

Uses reverse transcriptase, DNA polymerase, restriction enzymes, and ligase to assemble vector DNA.

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Gene Duplication

Gene duplication leads to similar genes that evolve independently, potentially acquiring different functions over time.

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Bacterial DNA Transfer

Conjugation involves direct DNA transfer, while transformation involves uptake of foreign DNA from the environment.

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Phosphorylation in Disease

Hyperphosphorylated proteins require increased phosphatase or decreased kinase activity to restore normal function.

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Ribosome Assembly

Ribosome assembly occurs in the nucleolus.

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Telomeres & Centromeres

Both telomeres and centromeres contain heterochromatin, which is inactive and tightly packed DNA.

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DNA Breaks & Repair

DSBs occur in the deoxyribose-phosphate backbone, requiring phosphodiester bond formation for repair.

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Wobble Base Pairing

The third codon position can vary without altering the encoded amino acid.

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Peptide Bond Formation

Translation forms peptide bonds through condensation, which releases water.

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Eukaryotic Ribosomes

Eukaryotic ribosomes are 80S, composed of 60S and 40S subunits, synthesizing proteins in the cytoplasm or rough ER.

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Rough ER & Secretory Proteins

Ribosomes attach to the rough ER for protein synthesis, rather than the Golgi apparatus.

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Antisense Oligonucleotides (ASOs)

Antisense oligonucleotides (ASOs) affect pre-mRNA splicing, changing exon inclusion and boosting functional protein production.

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MicroRNA

MicroRNAs (miRNAs) regulate gene expression by binding to complementary mRNA sequences, blocking translation or causing mRNA breakdown.

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Wild-type Organisms

Wild-type organisms have a fully functional, active gene, serving as the control.

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cDNA Structure

cDNA, made from mature mRNA lacks introns and doesn't undergo splicing but still produces the original protein.

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mRNA gel electrophoresis

Full-length mRNA (all exons included) migrates slower than shorter or truncated mRNA in gel electrophoresis.

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Timing of Splicing

Splicing occurs after transcription, before mRNA exits the nucleus, and involves the editing of pre-mRNA.

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Enzyme inhibition and modifications

Inhibiting enzymes that remove chemical groups (like acetyl groups) increases those modifications and gene expression.

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Histone Acetylation Increases Gene Expression

Increasing histone acetylation opens chromatin, increasing DNA accessibility and transcription.

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Study Notes

  • DNA polymerase replicates DNA, while RNA polymerase transcribes DNA into RNA.
  • DNA polymerase reads DNA in the 3′ → 5′ direction but synthesizes new strands in the 5′ → 3′ direction.
  • During DNA synthesis, the 3′ OH of the growing strand links to the 5′ phosphate of incoming nucleotides.
  • Each DNA replication cycle reduces the proportion of DNA helices containing original (15N) strands.
  • Double-strand breaks (DSBs) occur in the deoxyribose-phosphate backbone of DNA.
  • DSB repair necessitates the formation of phosphodiester bonds.
  • Mismatch Repair (MMR) corrects errors in DNA replication by replacing mismatched nucleotides.
  • MMR does not function on RNA.
  • DNA synthesis involves a condensation reaction, releasing water.
  • DNA synthesis is an exergonic reaction that releases energy through pyrophosphate hydrolysis.
  • Activators enhance transcription by aiding RNA polymerase binding, while repressors inhibit by blocking RNA polymerase.
  • RNA polymerase is essential for transcription but not for DNA replication itself.
  • To interpret RT-PCR results, reverse complement cDNA sequences and replace T with U to find the original mRNA sequence.
  • Alternative splicing allows one gene to produce multiple proteins.
  • Alternative splicing does not cause codon redundancy.
  • The poly-A tail (at the 3′ end) protects mRNA and facilitates nuclear export.
  • The 5′ cap prevents mRNA degradation and aids ribosome binding.
  • The genetic code is redundant (degenerate), where multiple codons can code for the same amino acid.
  • The genetic code is not ambiguous because each tRNA carries only one type of amino acid.
  • Wobble base pairing allows variation in the third codon position without altering the encoded amino acid.
  • Silent mutations do not change the amino acid sequence.
  • Missense mutations result in one amino acid being replaced by another.
  • Nonsense mutations create a premature stop codon.
  • Frameshift mutations occur from insertions or deletions that shift the reading frame.
  • Peptide bond formation during translation occurs through condensation.
  • rRNA forms ribosomes and catalyzes peptide bond formation.
  • Eukaryotic ribosomes are 80S, composed of 60S and 40S subunits, and synthesize proteins in the cytoplasm or rough ER.
  • Prokaryotic ribosomes are 70S (50S + 30S) and are not used in eukaryotic cells.
  • Ribosomes attach to the rough ER for synthesizing secretory proteins.
  • Cap-dependent translation decreases as cap analog concentration increases, while IRES-mediated translation is unaffected.
  • miRNAs silence genes by binding to complementary mRNA, which blocks translation or degrades the mRNA.
  • snRNAs splice pre-mRNA in the nucleus.
  • snRNAs are not involved in translation or gene silencing.
  • cDNA lacks introns and does not require splicing.
  • cDNA still produces the same protein as the original gene.
  • mRNA half-life analysis involves converting mRNA to cDNA and using qRT-PCR to track degradation.
  • Euchromatin (with acetylated histones) is open and active in transcription.
  • Heterochromatin (with deacetylated/methylated histones) is tightly packed and has low transcription.
  • Acetylation loosens chromatin, increasing transcription, while deacetylation condenses chromatin, decreasing transcription.
  • Somatic cells rarely express TERT, so the TERT gene is stored in heterochromatin.
  • In nonreducing SDS-PAGE, the homodimer migrates slower because of intact disulfide bonds, while the monomer migrates faster.
  • Northern blotting separates mRNA by size; smaller fragments move further down the gel.
  • Complementary nucleic acid strands hybridize, but proteins and antibodies do not.
  • Labeled RNA probes confirm gene expression by detecting mRNA.
  • Constructing vector DNA uses reverse transcriptase, DNA polymerase, restriction enzymes, and ligase.
  • DNA methylation occurs on cytosine (C) and reduces transcription.
  • Gene duplication creates similar genes that can mutate and evolve different functions.
  • Bacterial DNA transfer occurs through conjugation (direct transfer) and transformation (uptake of DNA from the environment).
  • Hyperphosphorylated proteins in diseases like Alzheimer's require increased phosphatase or decreased kinase activity to restore function.
  • Ribosome assembly occurs in the nucleolus.
  • Telomeres and centromeres both contain heterochromatin.
  • Telomeres shorten with each division and require telomerase.
  • Restriction enzymes recognize and cut palindromic sequences.
  • Telomeres (TTAGGG repeats) are not palindromes and are not cut by restriction enzymes
  • Full-length mRNA (with all exons) is the largest and migrates the least in gel electrophoresis.
  • Shorter or truncated mRNA (with exon removed) migrates further down.
  • Small RNAs (snRNA) are migrates the farthest down.
  • Splicing occurs after transcription is completed.
  • Splicing does not affect RNA synthesis.
  • Splicing occurs before mature mRNA leaves the nucleus.
  • Splicing does not control transport.
  • Splicing changes the mature mRNA sequence, indirectly affecting translation.
  • Inhibiting enzymes that remove chemical groups increases those modifications, typically boosting gene expression by loosening DNA structure.
  • Increasing histone acetylation generally increases gene expression by opening DNA structure.
  • Acetylation opens chromatin structure (euchromatin), increasing DNA accessibility and boosting gene transcription.
  • Deacetylation closes chromatin structure (heterochromatin), decreasing DNA accessibility and reducing gene transcription.
  • Antisense oligonucleotides (ASOs) bind to pre-mRNA and affect its splicing, increasing functional protein production.
  • MicroRNAs (miRNAs) regulate gene expression by binding complementary mRNA sequences, blocking translation or triggering mRNA breakdown.
  • snRNA/snRNPs edit (splice) pre-mRNA in the nucleus and are not involved in gene silencing or translation control.
  • Mutations in enhancers can increase transcription by allowing more transcription activators to bind, increasing gene expression.
  • Wild-type organisms have a fully functional, active gene and serve as a baseline or control.
  • Knockout organisms have an intentionally inactivated or nonfunctional gene to identify its biological role.
  • cDNA lacks introns and does not undergo splicing.
  • cDNA still produces the same protein as the endogenous gene.
  • cDNA expression levels can vary based on cloning factors, but the resulting protein remains unchanged.
  • Repressors inhibit transcription by preventing RNA polymerase from binding to the promoter.
  • Activators facilitate transcription by helping RNA polymerase bind more efficiently.
  • Genes that are rarely transcribed, are typically stored in heterochromatin.
  • The heterochromatin limits accessibility to RNA polymerase and reduces gene expression.

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Explore DNA replication and transcription processes. Learn about the roles of DNA and RNA polymerases, DNA synthesis directionality, and the importance of phosphodiester bonds in DNA repair. Understand mismatch repair mechanisms, energy dynamics, and the influence of activators and repressors on transcription.

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