Molecular Biology: Central Dogma, Gene Expression, and DNA Technologies

Central dogma of molecular biology

The framework describing the flow of genetic information from DNA to RNA to protein.

Gene expression

The process by which information from a gene is used to synthesize a functional gene product, typically a protein.

Modified central dogma

An updated version of the central dogma that includes processes such as reverse transcription.

Reverse transcriptase

An enzyme that synthesizes DNA from an RNA template.

Replicase/RNA-dependent RNA polymerase

An enzyme that replicates RNA from an RNA template.

Difference between retroviruses and RNA viruses

Retroviruses use reverse transcriptase to convert their RNA into DNA, while RNA viruses do not.

DNA reading (DNA sequencing)

The process of determining the precise order of nucleotides within a DNA molecule.

Human genome size

Approximately 3 billion base pairs.

Protein-coding genes in the human genome

Approximately 20,000 to 25,000 genes.

DNA writing (DNA synthesis)

The process of creating DNA molecules from nucleotides.

Synthetic genomes - Advances so far

The development of artificially created genomes for various applications.

Limitations in preparing synthetic genomes

Challenges include complexity, cost, and potential errors in synthesis.

Number of genes required to sustain life

A minimum of about 300 genes is estimated to be necessary.

DNA editing

The process of making specific changes to the DNA sequence of an organism.

CRISPR/Cas9

A revolutionary gene-editing technology that allows for precise modifications to DNA.

PAM sequence

A short DNA sequence that is essential for the CRISPR/Cas9 system to recognize and bind to the target DNA.

Non-Homologous End Joining (NHEJ)

A pathway that repairs double-strand breaks in DNA by directly joining the ends.

Homology Directed Repair (HDR)

A mechanism that repairs double-strand breaks in DNA using a homologous template.

Applications of CRISPR/deadCas9 in molecular genetics

Used for gene regulation and epigenetic modifications without altering the DNA sequence.

CRISPR/Cas system to degrade RNA

Utilizes CRISPR technology to target and degrade RNA molecules.

Cas13

A CRISPR-associated enzyme that targets RNA for degradation.

Dead cas9

A modified version of Cas9 that can bind to DNA but does not cut it, used for gene regulation.

Dead cas13

A modified version of Cas13 that can bind to RNA but does not degrade it, used for RNA regulation.

Prime editing

A precise method for editing genes that allows for targeted insertions, deletions, and replacements.

S and R strains of Streptococcus pneumoniae

Different strains of bacteria where S is smooth (pathogenic) and R is rough (non-pathogenic).

Pathogenic and nonpathogenic strains of bacteria

Pathogenic strains cause disease while nonpathogenic strains do not.

Transformation

The process by which a cell takes up foreign DNA from its environment.

DNase

An enzyme that degrades DNA.

RNase

An enzyme that degrades RNA.

Protease

An enzyme that degrades proteins.

Bacteriophage

A virus that infects and replicates within bacteria.

Labeling of DNA and protein

Techniques used to tag DNA or proteins for visualization or tracking.

Purines

A type of nitrogenous base that includes adenine and guanine.

Pyrimidines

A type of nitrogenous base that includes cytosine, thymine, and uracil.

Polymer

A large molecule composed of repeating structural units (monomers).

Pentose sugar

A five-carbon sugar molecule, such as ribose or deoxyribose.

Ribose sugar

A five-carbon sugar present in RNA.

Deoxyribose sugar

A five-carbon sugar present in DNA.

Nucleosides

Molecules consisting of a nitrogenous base and a sugar.

Nucleotides

The basic building blocks of nucleic acids, consisting of a nucleoside and a phosphate group.

Ribonucleotides

Nucleotides that contain ribose sugar.

Deoxyribonucleotides

Nucleotides that contain deoxyribose sugar.

Phosphodiester bond

The bond that links nucleotides together in a nucleic acid.

Glycosidic bond

The bond that connects a sugar to a nitrogenous base in nucleotides.

Antiparallel

Describes the opposite orientation of the two strands of DNA.

AT/GC rule

The principle that adenine pairs with thymine and guanine pairs with cytosine in DNA.

Backbone

The sugar-phosphate structure that forms the structural framework of nucleic acids.

B DNA

The most common form of DNA in cells, characterized by a right-handed helix.

X-ray crystallography

A technique used to determine the three-dimensional structure of molecules.

Chargaff's rule

The observation that in DNA, the amount of adenine equals thymine and the amount of guanine equals cytosine.

Complementary

Describes the relationship between nucleotide bases that pair with each other in DNA.

A DNA

A form of DNA that is more compact than B DNA and has a right-handed helix.

Z DNA

A form of DNA that has a left-handed helix and is thought to play a role in gene expression.

DNA denaturation

The process of separating the two strands of DNA.

Melting temperature (Tm)

The temperature at which half of the DNA strands are in the double-helix state and half are in the 'melted' single-stranded state.

DNA renaturation

The process of re-forming the double helix structure of DNA after denaturation.

Semiconservative mode of replication

The process by which DNA replicates, producing two copies that each contain one original strand and one new strand.

Density gradient centrifugation

A technique used to separate molecules based on their density by spinning them at high speeds in a gradient of a dense medium.

Conservative mode of replication

A hypothetical model of DNA replication where the original DNA molecule remains intact and a completely new molecule is synthesized.

Dispersive mode replication

A model of DNA replication where the parental DNA is dispersed into two new molecules, each containing segments of old and new DNA.

Origin of replication in bacteria - OriC

The specific location on a bacterial chromosome where DNA replication begins.

Bidirectional DNA synthesis

A method of DNA replication where synthesis occurs in two directions from a single origin of replication.

Replication fork

The Y-shaped region where the DNA double helix is unwound and replicated.

Template strands

The original strands of DNA that serve as a guide for the synthesis of new complementary strands.

DNA polymerases

Enzymes that synthesize new DNA strands by adding nucleotides to a growing chain.

Leading and lagging strands

The two strands of DNA during replication; the leading strand is synthesized continuously, while the lagging strand is synthesized in short fragments.

Okazaki fragments

Short segments of DNA synthesized on the lagging strand during DNA replication.

Initiator proteins

Proteins that bind to the origin of replication and initiate the process of DNA replication.

DnaA box

A specific DNA sequence recognized by DnaA proteins to initiate DNA replication in bacteria.

DnaA proteins

Proteins that bind to the DnaA box and facilitate the unwinding of DNA at the origin of replication.

Primase

An enzyme that synthesizes short RNA primers needed for DNA polymerase to start DNA synthesis.

Helicase

An enzyme that unwinds the DNA double helix at the replication fork.

Primer

A short strand of RNA or DNA that provides a starting point for DNA synthesis.

DNA ligase

An enzyme that joins Okazaki fragments together by forming phosphodiester bonds.

Exonuclease activity of DNA polymerase

The ability of DNA polymerase to remove nucleotides from the ends of a DNA strand, allowing for proofreading and correction.

Single strand DNA binding proteins

Proteins that stabilize single-stranded DNA during replication to prevent it from re-annealing.

DNA proofreading

The process by which DNA polymerases check and correct errors during DNA replication.

DNA repair mechanism - Mismatch repair

A system that corrects errors that occur during DNA replication, specifically mismatched base pairs.

Bacterial origin of replication

Key features include specific sequences where DNA replication begins.

DnaA protein

Initiates DNA replication by binding to the origin of replication and unwinding the DNA.

Lagging strand synthesis

Synthesized in fragments due to the antiparallel nature of DNA and the direction of DNA polymerase activity.

Enzymes at the replication fork

Various enzymes and proteins function to synthesize new strands of DNA, including helicase, primase, and DNA polymerases.

Leading vs. lagging strands

Leading strand is synthesized continuously, while lagging strand is synthesized in Okazaki fragments.

Proofreading function of DNA polymerase

DNA polymerase has the ability to correct errors during DNA synthesis, ensuring fidelity.

Origins of replication in bacteria vs. eukaryotes

Bacteria typically have a single origin of replication, while eukaryotes have multiple origins.

Functions of DNA polymerases in eukaryotes

Different DNA polymerases are involved in DNA replication, repair, and mitochondrial DNA synthesis.

DNA replication at eukaryotic chromosome ends

Occurs through mechanisms such as telomerase activity to prevent loss of genetic information.

Consequences of mutations in DNA repair enzymes

Mutations in enzymes like those causing Xeroderma pigmentosum, MutS, and MutL, and BRCA can lead to increased cancer risk.

Double stranded DNA break model of recombination

Involves steps such as break formation, strand invasion, and resolution, with alternative pathways for resolution.

Gene conversion during recombination

Occurs when genetic information is transferred from one DNA molecule to another during homologous recombination.

RNA as an intermediate molecule

Evidence includes the role of RNA in transcription and translation processes in gene expression.

Organization of a protein-encoding gene

Includes regulatory elements, coding sequences, and introns/exons, which are transcribed into mRNA.

Stages of transcription in prokaryotes

Include initiation, elongation, and termination.

Characteristics of a bacterial promoter

Includes specific sequences recognized by RNA polymerase and transcription factors.

RNA polymerase transcribing a bacterial gene

RNA polymerase binds to the promoter and synthesizes RNA complementary to the DNA template.

Consensus sequences in prokaryotic promoters

Common sequences found in bacterial promoters that are recognized by RNA polymerase.

Transcriptional termination mechanisms in bacteria

Include Rho-dependent and Rho-independent termination.

Functions of RNA polymerases in eukaryotes

Include RNA polymerase I for rRNA, RNA polymerase II for mRNA, and RNA polymerase III for tRNA and other small RNAs.

Characteristics of a eukaryotic promoter

Includes TATA box and other regulatory elements that facilitate transcription initiation.

Assembly of general transcription factors and RNA polymerase

Occurs at the promoter of eukaryotic genes to initiate transcription.