Unit notes

DNA sequence can be transcribed into RNA, and an mRNA coding sequence can be translated into protein. A codon is a three-base mRNA sequence; an anticodon is a complementary sequence on tRNA. DNA polymerase, RNA polymerase, and ribosomes perform different jobs.

DNA strands are antiparallel, and complementary base pairing allows each parental strand to serve as a template. Semiconservative replication produces molecules with one old and one newly synthesized strand. DNA polymerases extend from an existing 3′ end, so new DNA is synthesized 5′ to 3′.

A replication fork produces a leading strand and a lagging strand because the two templates have opposite orientations. The lagging strand is synthesized in fragments that are later joined. Proofreading and repair reduce errors but do not eliminate mutation. A mutation must occur in a lineage contributing to gametes to be inherited by offspring in a sexually reproducing animal.

RNA polymerase uses a DNA template strand to synthesize complementary RNA. The RNA sequence corresponds to the coding strand except that uracil replaces thymine. Identify strand direction before converting a sequence; copying the template letters without complementing them gives the wrong RNA.

In eukaryotes, pre-mRNA commonly receives a 5′ cap and poly-A tail and undergoes splicing. Alternative splicing can generate different mature transcripts from one gene. Introns are removed from the RNA, not cut out of the cell’s genomic DNA each time the gene is expressed. Regulation can occur at transcription, RNA processing, RNA stability, translation, or protein modification.

Ribosomes read mRNA codons in the 5′ to 3′ direction. Transfer RNAs pair anticodons with codons and deliver amino acids. The start codon establishes a reading frame; stop codons recruit termination machinery rather than specifying ordinary amino acids. Protein synthesis proceeds from the amino end toward the carboxyl end.

A base substitution can be synonymous, missense, or nonsense depending on its position and resulting codon. An insertion or deletion not divisible by three can shift a coding reading frame. Effects depend on location: a regulatory mutation can change expression without changing the protein’s coding sequence.

In bacteria, operon regulation coordinates genes with related functions. Repressors, activators, and small regulatory molecules influence transcription. An inducible system can increase expression when a relevant substrate is available; a repressible system can reduce expression when a product is abundant. Always reason from the specific regulatory interactions provided.

In eukaryotes, transcription factors bind regulatory DNA and interact with transcription machinery. Chromatin state affects access to DNA. Cells can contain the same genome yet express different sets of genes. Epigenetic regulation changes gene activity without requiring a change in nucleotide sequence, but a claim of transgenerational inheritance requires specific evidence.

PCR amplifies a selected DNA region using primers and repeated temperature cycles. Gel electrophoresis separates many DNA fragments primarily by size, with smaller fragments generally moving farther through the gel. A matching band size is evidence about length, not proof of identical full sequence.

A reporter gene can reveal effects of a regulatory sequence. Compare matched constructs that differ in the tested feature, normalize relevant quantities, and include positive and negative controls. Gene editing can disrupt or alter a target, but off-target effects and delivery success may require checks before assigning every observed phenotype to the intended change.

DNA polymerase adds nucleotides to a growing strand’s 3′ end, so synthesis proceeds 5′ to 3′. Because the two templates are antiparallel, one new strand can be synthesized continuously toward a replication fork while the other is assembled in fragments. Primase provides starting points and ligase joins DNA fragments. Both new strands still grow by the same chemical direction of nucleotide addition.

RNA polymerase reads a DNA template 3′ to 5′ while producing RNA 5′ to 3′. The coding DNA strand has the same base order as the RNA except that DNA uses T and RNA uses U. Identify which strand and direction are provided before writing an RNA sequence. Eukaryotic RNA processing includes removal of introns and joining of exons; alternative splicing can generate different mature messages from one primary transcript.

A base substitution can be silent, change an amino acid, or introduce a stop codon, depending on its position and the genetic code. An insertion or deletion in a coding region can shift the reading frame if its size is not a multiple of three. The same change outside a coding region may instead affect regulation or RNA processing. Start by locating the mutation rather than assuming every mutation changes every downstream amino acid.

Even an amino-acid change does not automatically destroy function. Compare the properties of the original and replacement amino acids, the location in the protein, and the measured phenotype. A change near a binding site can have a different effect from a change in an exposed flexible region. Regulatory mutations can alter how much protein is made or which cells make it without changing the protein’s amino-acid sequence.

PCR amplifies a selected DNA region using primers, a thermostable polymerase, and repeated temperature changes. The primers determine which region is amplified. A negative control without template helps detect contamination; a positive control helps establish that the reagents and conditions can work. A missing product can reflect absent target DNA or a failed reaction, so controls matter.

Gel electrophoresis separates DNA fragments largely by size in a standard agarose gel; smaller fragments generally travel farther. A band’s position estimates fragment length relative to a size standard, not the identity of every base. A gene-expression comparison might measure RNA abundance, protein abundance, or activity. Those are related but not identical: translation efficiency and protein degradation can separate RNA levels from functional protein levels.

A gene is not simply a stretch that a ribosome reads directly from DNA. A promoter helps determine where transcription begins; regulatory sequences influence how often it begins. A transcribed region can include untranslated regions and, in eukaryotes, introns that are removed. Only the translated coding region determines the amino-acid sequence. A mutation can therefore change expression without changing the encoded protein, or change the protein while leaving the amount of transcript approximately unchanged.

Separate the measurements in a data table. DNA quantity can estimate copy number; RNA abundance reflects both transcription and RNA breakdown; protein abundance reflects both translation and protein breakdown. Enzyme activity additionally depends on folding, localization, modification and environmental conditions. If RNA rises but activity does not, that observation narrows possible explanations but does not by itself identify one. Measuring protein abundance and activity per unit protein helps distinguish a production problem from a functional problem.

In an explanation, name the affected step and connect it to the output. “The mutation affects the gene” is much weaker than “the altered promoter reduces polymerase recruitment, decreasing transcript production and therefore the template available for translation.” Preserve uncertainty when the experiment measured only the final output.

Write 5′ and 3′ labels before you complement a strand. Bases pair antiparallel, and both DNA and RNA polymerases extend a new strand at its 3′ end. A template written 3′ to 5′ can be complemented left to right into a product written 5′ to 3′. If the template is supplied 5′ to 3′, first determine the complementary strand and then reverse its written order when a 5′-to-3′ answer is required. Direction is a property of the molecule, not of where a diagram places the fork.

Translation reads the mature mRNA in three-base groups beginning in the relevant reading frame. A substitution can be synonymous, missense or nonsense depending on the codon change. An insertion or deletion in a coding region shifts the frame when its length is not a multiple of three. A three-base deletion can remove an amino acid without shifting all later groups, but it can still disrupt an essential binding region. A mutation outside the coding region can alter splicing or regulation.

Do not infer a protein’s exact length without locating its stop codon in the new frame. Likewise, a changed amino acid does not automatically mean complete loss of function. Its chemical properties, location and interactions determine the effect; experimental activity data are stronger evidence than the mere fact that a sequence changed.

In a bacterial regulatory model, a repressor that binds an operator can block transcription. A small molecule can alter the repressor’s binding, linking a nutrient condition to expression. Determine whether the molecule turns binding on or off in the supplied model; the word “regulator” alone does not tell you whether transcription increases. A mutation that prevents repressor binding may allow transcription even under conditions that normally repress it.

Eukaryotic control includes chromatin accessibility, transcription-factor binding, RNA processing, RNA stability, translation and protein modification. Different cell types can share a genome while expressing different sets of genes. A signal may activate a transcription factor, which changes expression, which later changes a cellular phenotype. This sequence also explains why a response involving newly made protein may take longer than changing the activity of protein already present.

To evaluate a regulatory hypothesis, compare appropriate controls at the step in question. A promoter reporter tests regulatory output in the reporter’s context. Restoring a functional regulator can support causation, especially if it reverses the phenotype. It does not automatically exclude every indirect effect. Keep the conclusion proportional to the experiment.

PCR amplifies a chosen DNA region using primers that flank it. A positive result supports the presence of an amplifiable target; it does not automatically show that the gene was transcribed or functional. A negative control without template helps detect contamination, while a positive control helps distinguish target absence from failure of the reaction. Primer binding and sample quality are additional possible explanations for a missing band.

Gel electrophoresis separates DNA fragments mainly by size under comparable conditions: smaller fragments generally travel farther. Compare a lane with the ladder, not with an arbitrary distance from the page margin. A band at the predicted size supports a size claim; many sequences can have that size. Restriction patterns or sequencing can provide different kinds of evidence.

For expression experiments, compare amounts after suitable normalization. More signal may reflect more cells, more starting RNA, or greater expression per cell. A reference measurement helps address these alternatives but is useful only if it remains sufficiently stable under the tested conditions. Explain what the control rules out rather than calling it “something to compare with.”