Genetics Test 2: DNA/RNA/Mutations

1. DNA and RNA structure

DNA (deoxyribonucleic acid)

A polymer and macromolecule that stores the genetic instructions needed for an organism to develop, function, and reproduce. Example: DNA contains genes whose sequences can be transcribed into RNA; DNA must also be replicated before a cell divides so each daughter cell can receive genetic information.

RNA (ribonucleic acid)

A nucleic acid that helps use genetic information and can also perform regulatory or structural functions. Example: Messenger RNA (mRNA) carries a copy of information from DNA to the machinery that makes proteins. Transfer RNA (tRNA) helps deliver amino acids during translation, and ribosomal RNA (rRNA) is part of the ribosome.

Genome

The complete set of genetic material in a cell or organism. Example: The human genome includes DNA from chromosomes, including protein-coding genes, regulatory regions, and repetitive sequences.

Nucleotide

The building block, or monomer, of DNA and RNA. It contains a nitrogenous base, a five-carbon sugar, and phosphate group(s). Example: A DNA nucleotide containing adenine has deoxyribose and a phosphate group as well as adenine.

Nitrogenous base

The information-carrying component of a nucleotide; the sequence of bases varies along a nucleic acid strand. Example: Changing a DNA base from A to G changes the sequence and may affect a gene's product.

Purines

The two-ring nitrogenous bases adenine (A) and guanine (G). Example: Depurination removes adenine or guanine from DNA.

Pyrimidines

The one-ring nitrogenous bases cytosine (C), thymine (T), and uracil (U). Example: DNA contains cytosine and thymine, while RNA uses uracil instead of thymine.

Deoxyribose

The five-carbon sugar found in DNA. Example: Deoxyribose forms part of the DNA sugar-phosphate backbone and differs from ribose by having one less oxygen atom.

Ribose

The five-carbon sugar found in RNA. Example: RNA nucleotides contain ribose, which contributes to RNA's chemical properties and generally lower stability compared with DNA.

Polymer

A large molecule made of repeating smaller units called monomers. Example: DNA is a polymer made from nucleotide monomers joined into a strand.

Sugar-phosphate backbone

The repeating sugar and phosphate structure that forms the outside of a nucleic acid strand; the bases project from it. Example: DNA bases can pair in the center of the double helix while the sugar-phosphate backbones form its outer structure.

Phosphodiester bond

A covalent bond connecting the 3′ oxygen of one nucleotide's sugar to the 5′ phosphate of the next nucleotide. Example: When a new nucleotide is added to a growing DNA strand, a phosphodiester bond connects it to the existing strand.

5′ end and 3′ end

The two chemically distinct ends of a nucleic acid strand, defined by the numbered carbons of its sugar. Example: A DNA strand may be written 5′-ATGC-3′; DNA polymerase adds a nucleotide to the growing strand's 3′ end.

Directionality

The orientation of a nucleic acid strand from 5′ to 3′. Example: DNA sequences are conventionally written 5′ to 3′, and DNA polymerase synthesizes a new strand in that direction.

Complementary base pairing

Specific pairing between bases: A pairs with T in DNA, A pairs with U in RNA, and G pairs with C. Example: The DNA strand 5′-ATGC-3′ has the complementary strand 3′-TACG-5′.

Chargaff’s rule

In double-stranded DNA, the amount of adenine equals thymine and the amount of guanine equals cytosine. Example: If a double-stranded DNA sample contains 20% A, it contains 20% T; the remaining 60% is split into 30% G and 30% C.

Hydrogen bond

A noncovalent interaction that helps hold complementary bases together. Example: A–T pairs form two hydrogen bonds, while G–C pairs form three.

Antiparallel strands

The two DNA strands run in opposite directions: one 5′ to 3′ and the other 3′ to 5′. Example: If one strand is written 5′-AAGC-3′, its complementary strand is 3′-TTCG-5′.

Double helix

The twisted, two-stranded structure of DNA. Example: The strands of DNA wind around one another, with complementary bases inside and sugar-phosphate backbones outside.

Base stacking

Stabilizing interactions between neighboring bases along a nucleic acid strand. Example: Bases stack on top of neighboring bases inside the DNA helix, contributing to DNA stability in addition to hydrogen bonding.

Major groove and minor groove

The larger and smaller grooves along the surface of the DNA double helix. Example: Proteins can recognize specific DNA sequences by interacting with chemical features exposed in the grooves.

B-DNA

The common right-handed DNA helix found under typical physiological conditions, with roughly 10–10.5 base pairs per turn. Example: Most textbook depictions of cellular DNA show B-DNA.

Z-DNA

A less common, left-handed DNA helix with a zigzag-shaped backbone that can form under particular conditions, including certain sequences and supercoiling stress. Example: Z-DNA can form in regions of DNA under torsional stress and may be involved in gene regulation.

DNA methylation

The addition of methyl groups to DNA bases; it can influence DNA organization and gene regulation. Example: Methylation of cytosine at certain sites can affect how genes are expressed.

RNA secondary structure

Local folding caused by base pairing within a single RNA molecule. Example: Complementary regions of an RNA strand can pair with each other to form stems, loops, and bulges.

RNA tertiary structure

The overall three-dimensional folding of an RNA molecule, influenced by interactions among distant regions, ions, small molecules, and proteins. Example: A folded tRNA adopts a three-dimensional shape that supports its role in translation.

Messenger RNA (mRNA)

RNA that carries information copied from DNA and can be used to direct protein synthesis. Example: A gene is transcribed into mRNA, which is then read in codons during translation.

Transfer RNA (tRNA)

RNA that carries amino acids to the ribosome and recognizes mRNA codons through its anticodon. Example: A tRNA with an appropriate anticodon pairs with an mRNA codon and delivers the corresponding amino acid.

Ribosomal RNA (rRNA)

RNA that forms part of ribosomes and helps them carry out protein synthesis. Example: rRNA contributes to the structure and catalytic activity of the ribosome.

Central dogma

The general flow of genetic information from DNA to RNA to protein. Example: A gene's DNA sequence can be transcribed into mRNA and the mRNA can be translated into a protein.

2. Experiments establishing DNA as genetic material

Genetic material

Material that stores biological information, can be passed between generations, can be replicated, and can vary. Example: DNA meets these requirements because it carries instructions, can be inherited and copied, and can acquire mutations.

Transformation

A process in which a cell takes up genetic material and acquires a new trait. Example: In Griffith's experiment, live rough bacteria acquired the ability to produce the smooth, disease-causing type after exposure to material from heat-killed smooth bacteria.

Transforming principle

The substance that Griffith inferred could transfer information from one bacterial strain to another; later work established that DNA was responsible. Example: Material from dead smooth bacteria transformed live rough bacteria into smooth bacteria.

Frederick Griffith’s experiment

An experiment showing that material from heat-killed smooth bacteria could transform live rough bacteria. Example: Live rough bacteria alone did not kill the mice, and heat-killed smooth bacteria alone did not kill them, but the mixture of live rough and heat-killed smooth bacteria resulted in live smooth bacteria being recovered from the mice.

Avery, MacLeod, and McCarty experiment

An experiment identifying DNA as the transforming substance by testing extracts treated with enzymes that destroyed different types of molecules. Example: Transformation continued after RNA or proteins were destroyed but stopped when DNA was destroyed by DNase.

DNase

An enzyme that breaks down DNA. Example: In the Avery–MacLeod–McCarty experiment, DNase treatment prevented transformation.

RNase

An enzyme that breaks down RNA. Example: Treating a bacterial extract with RNase did not eliminate its transforming ability in the Avery–MacLeod–McCarty experiment.

Protease

An enzyme that breaks down proteins. Example: Protease treatment did not prevent transformation in the Avery–MacLeod–McCarty experiment, supporting the conclusion that protein was not the transforming principle.

Hershey–Chase experiment

An experiment using bacteriophages to show that DNA, rather than protein, enters bacterial cells and carries the genetic information needed to produce new phages. Example: Radioactively labeled phage DNA entered bacterial cells, while most labeled phage protein remained outside.

Rosalind Franklin and Maurice Wilkins

Scientists whose X-ray diffraction studies provided evidence about DNA's helical structure. Example: Franklin's diffraction data supported a helical arrangement and helped reveal structural features of DNA.

Linus Pauling

A scientist whose work on molecular structure and model building influenced approaches to understanding biological molecules. Example: His ball-and-stick modeling work helped establish the value of constructing molecular models to explain structure.

Erwin Chargaff

A scientist who measured DNA base composition and identified the relationships later expressed as Chargaff's rule. Example: His findings showed that the amount of A generally matched T and G matched C in double-stranded DNA.

James Watson and Francis Crick

Scientists who proposed the double-helical model of DNA in 1953, consistent with existing evidence. Example: Their model incorporated complementary base pairing, antiparallel strands, and the DNA helix.

3. DNA replication

DNA replication

The process of copying DNA before cell division. Example: Before a cell divides, its DNA is replicated so that the daughter cells can receive genetic information.

Semiconservative replication

Replication in which each daughter DNA double helix contains one original strand and one newly synthesized strand. Example: After one round of replication, each DNA molecule contains one parental strand paired with one new strand.

Conservative replication model

A proposed model in which the original two strands remain together and an entirely new double helix is made. Example: The parental DNA molecule would remain intact while the second molecule contained two new strands; this is not the accepted model of DNA replication.

Dispersive replication model

A proposed model in which parental and newly synthesized DNA would be interspersed within both strands. Example: Each daughter strand would contain a mixture of old and new DNA segments; this is not the accepted model.

Meselson–Stahl experiment

An experiment using different nitrogen isotopes to distinguish parental DNA from newly synthesized DNA and test replication models. Example: After one round of replication, DNA showed intermediate density, supporting semiconservative replication.

Origin of replication

A DNA sequence or region where replication begins. Example: Bacteria typically use a particular origin, while eukaryotic chromosomes have multiple origins.

Bidirectional replication

Replication proceeding away from an origin in two directions. Example: Two replication forks move outward from a single origin along the DNA.

Replication fork

The region where DNA strands separate and new DNA strands are synthesized. Example: Helicase unwinds the parental DNA at the fork, allowing polymerases to copy the templates.

Helicase

An enzyme that unwinds double-stranded DNA by separating the strands. Example: Helicase opens the double helix at a replication fork.

Single-strand binding proteins (SSBs)

Proteins that bind separated DNA strands and help prevent them from re-pairing. Example: SSBs stabilize the exposed templates after helicase unwinds DNA.

Primase

An enzyme that synthesizes a short RNA primer. Example: Primase makes an RNA primer that provides a free 3′ end for DNA polymerase to extend.

RNA primer

A short RNA segment that provides a starting point for DNA synthesis. Example: DNA polymerase cannot begin a new strand from nothing, so it extends from the primer's 3′ end.

DNA polymerase

An enzyme that synthesizes DNA using a template and adds nucleotides to a growing strand's 3′ end. Example: A DNA polymerase reads the template and adds complementary nucleotides to the new strand.

DNA polymerase III

The principal bacterial replicative DNA polymerase in the standard bacterial replication model. Example: It synthesizes most of the new DNA on the leading and lagging strands.

DNA polymerase I

A bacterial enzyme that removes RNA primers and replaces them with DNA during replication. Example: After an Okazaki fragment has been synthesized, DNA polymerase I removes the RNA primer and fills the resulting gap with DNA.

Leading strand

The new DNA strand synthesized continuously in the same overall direction that the replication fork moves. Example: DNA polymerase extends the leading strand continuously from a primer.

Lagging strand

The new DNA strand synthesized discontinuously in short segments because DNA polymerase can synthesize only in the 5′ to 3′ direction. Example: The lagging strand is built as multiple Okazaki fragments.

Okazaki fragment

A short DNA segment synthesized on the lagging strand. Example: Each fragment begins at an RNA primer, is extended by DNA polymerase, and is later joined to neighboring fragments.

DNA ligase

An enzyme that seals breaks in the sugar-phosphate backbone. Example: Ligase joins adjacent Okazaki fragments after their RNA primers have been replaced with DNA.

Replisome

The coordinated group of proteins that carries out DNA replication at a replication fork. Example: The replisome includes proteins responsible for unwinding DNA, priming, and DNA synthesis.

Primosome

A group of proteins involved in initiating DNA synthesis, including the production of primers. Example: Primase works with other replication proteins to make primers needed to begin DNA synthesis.

Processivity

The ability of an enzyme to perform repeated catalytic steps without dissociating from its substrate. Example: A highly processive DNA polymerase can add many nucleotides before leaving the DNA template.

Fidelity

The accuracy with which DNA is copied. Example: DNA polymerase has high fidelity because incorrect nucleotide incorporation is uncommon and proofreading can remove some mistakes.

Proofreading

A DNA polymerase activity that removes an incorrectly incorporated nucleotide, commonly through 3′ to 5′ exonuclease activity. Example: If polymerase inserts the wrong base, it can remove the mismatch and then add the correct nucleotide.

Topoisomerase

An enzyme that relieves DNA twisting and supercoiling by temporarily cutting and rejoining DNA. Example: As helicase unwinds DNA, topoisomerase helps prevent excessive twisting ahead of the replication fork.

DnaA

A bacterial protein involved in recognizing and initiating replication at the bacterial origin. Example: DnaA binds the E. coli origin, oriC, during initiation.

oriC

The origin of replication in E. coli. Example: Replication begins at oriC and proceeds in two directions.

Origin recognition complex (ORC)

A eukaryotic protein complex that recognizes replication origins and helps recruit replication-initiation machinery. Example: ORC binds an origin region before DNA replication begins.

MCM2–7 complex

A eukaryotic protein complex that functions as the replicative helicase. Example: Once activated, the MCM complex unwinds DNA during replication.

Cdc6 and Cdt1

Eukaryotic proteins involved in loading the MCM helicase during replication licensing. Example: They help load MCM2–7 onto DNA during preparation for replication.

Autonomously replicating sequence (ARS)

A DNA sequence that can function as a replication origin in yeast. Example: An ARS can allow a suitable DNA molecule to replicate in yeast cells.

Telomere

A repetitive DNA region at the end of a eukaryotic chromosome that helps protect chromosome ends. Example: Telomeres help prevent chromosome ends from being treated like broken DNA.

Telomerase

An enzyme complex that extends telomeres using its own RNA template. Example: Telomerase adds telomeric repeats to chromosome ends in cells that express sufficient telomerase activity.

TERT

The catalytic protein component of telomerase, called telomerase reverse transcriptase. Example: TERT helps synthesize additional telomeric DNA using telomerase's RNA component as a template.

Cellular senescence

A state in which a cell stops dividing. Several signals can cause senescence, including critically short telomeres. Example: A cell with severely shortened telomeres may stop dividing rather than continue with unstable chromosome ends.

4. PCR and DNA sequencing

Polymerase chain reaction (PCR)

A laboratory technique that amplifies a selected DNA region through repeated cycles of heating and cooling. Example: PCR can produce many copies of a DNA region from a small starting sample.

DNA template

The DNA molecule containing the sequence to be copied. Example: In PCR, the target region within the template is copied using primers and DNA polymerase.

Forward primer

A short DNA oligonucleotide that binds one side of the target region and provides a starting point for DNA synthesis. Example: The forward primer binds one template strand so polymerase can extend a new strand toward the target region.

Reverse primer

A short DNA oligonucleotide that binds the opposite strand at the other end of the target region. Example: The reverse primer enables synthesis from the other end so the DNA region between the two primers can be amplified.

Primer design

Choosing primers that bind the intended target region and have suitable sequence properties. Example: Primers should bind the intended sequences, avoid problematic self-pairing, and have compatible melting temperatures.

Primer length

The number of nucleotides in a primer. Example: A primer that is 20 nucleotides long contains 20 bases; length affects binding specificity and melting temperature.

Melting temperature ((T_m))

The temperature at which approximately half of a DNA duplex is separated into single strands under specified conditions. Example: If one primer has a higher (T_m) than another, the pair may require careful annealing-temperature selection.

Annealing temperature

The temperature during PCR at which primers bind to complementary template sequences. Example: If the temperature is too low, primers may bind to unintended sequences; if it is too high, primers may not bind efficiently.

Denaturation

The PCR step in which heating separates double-stranded DNA into single strands. Example: Heating to approximately 95°C separates the DNA strands before primer binding.

Annealing

The PCR step in which cooling allows primers to bind to complementary DNA sequences. Example: During the annealing step, forward and reverse primers bind on opposite strands at the target's boundaries.

Extension

The PCR step in which DNA polymerase extends primers by adding complementary nucleotides. Example: A thermostable polymerase extends the primers to make new DNA copies.

Thermostable DNA polymerase

A DNA polymerase that remains functional after the high temperatures used during PCR. Example: Taq polymerase can withstand repeated denaturation cycles.

Phusion high-fidelity polymerase

A high-fidelity DNA polymerase used to amplify DNA with a lower error rate than many standard PCR enzymes. Example: Phusion can be selected when accurate amplification is important for downstream sequencing or cloning.

dNTPs (deoxynucleoside triphosphates)

The DNA building blocks used by DNA polymerase during synthesis: dATP, dTTP, dCTP, and dGTP. Example: PCR requires all four dNTPs to copy a DNA target containing all four bases.

Magnesium ions ((\mathrm{Mg}^{2+}))

Ions required as cofactors for many DNA polymerases. Example: If magnesium concentration is unsuitable, PCR yield, specificity, and enzyme performance can change.

PCR cycle

One sequence of denaturation, annealing, and extension. Example: Repeating PCR cycles allows the target region to accumulate in large quantities.

Amplicon

The DNA product generated by PCR. Example: If primers flank a 400-base-pair region, the intended amplicon is approximately 400 base pairs long.

Target sequence

The particular DNA region intended for amplification or analysis. Example: Primers designed on opposite sides of a gene segment define the target sequence to be amplified.