Biotechnology and Bioinformatics Lecture Review

Comprehensive review practice flashcards for biotechnology and bioinformatics, covering cell structure, molecular genetics, recombinant DNA technologies, CRISPR/Cas9 systems, protein expression hosts, and regulatory ethics.

How is biotechnology defined in the lecture?

The production of goods and services using biological organisms, systems, and processes.

What is molecular biotechnology and what core technology does it use?

A field that uses recombinant DNA technology to manipulate DNA, including identifying and cloning genes, studying cloned gene expression, and producing large quantities of recombinant proteins.

What five major fields of modern biotechnology are represented by color codes in the lecture diagram?

Blue Biotechnology (marine and aquatic), Green Biotechnology (agriculture and food), Red Biotechnology (medical), White Biotechnology (industrial), and Yellow Biotechnology (insects).

What are examples of applications in Blue Biotechnology?

Development of pharmaceuticals from marine organisms, such as anti-cancer compounds from marine sponges, or algae-based biofuels.

What are examples of applications in Green Biotechnology?

Genetically modified (GM) crops such as pest-resistant Bt corn or drought-tolerant plants.

What are examples of applications in Red Biotechnology?

Production of insulin using genetically modified bacteria and development of gene therapy techniques for inherited diseases.

What are examples of applications in White Biotechnology?

Industrial enzyme applications used as catalysts to break down hazardous chemicals, manufacture bioethanol from agricultural waste, or produce bioplastics via microbial fermentation in bioreactors.

What are examples of applications in Yellow Biotechnology?

Using insects such as silkworms to produce silk for medical applications or utilizing insect-derived compounds to develop new antibiotics.

What is the basic and smallest unit of life?

The cell.

What are the four major classes of biomolecules and their primary functions described in the lecture?

Carbohydrates provide material to build the cell membrane; proteins provide structure and immunity; nucleic acids store genetic information; lipids store energy and provide material for the cell membrane.

What are the three evolutionary domains of life?

Bacteria, Archaea, and Eukaryotes.

Based on this comparative table, what are the primary structural differences between eukaryotes and prokaryotes?

Eukaryotes possess a nucleus, membrane-bound organelles with specific functions, linear DNA associated with histones, sizes of $10\text{--}100\,\mu\text{m}$, and can be unicellular or multicellular; prokaryotes lack a nucleus and membrane-bound organelles, have naked circular DNA in a nucleoid, measure $1\text{--}5\,\mu\text{m}$, and are always unicellular.

What are the typical size ranges of eukaryotic cells versus prokaryotic cells?

Eukaryotic cells are larger ($10\text{--}100\,\mu\text{m}$), whereas prokaryotic cells are smaller ($1\text{--}5\,\mu\text{m}$).

Why are ribosomes not classified as cellular organelles?

Because organelles are defined by having a surrounding membrane, which ribosomes lack.

What does the Endosymbiotic Theory explain regarding mitochondria and chloroplasts?

It explains that mitochondria and chloroplasts originated when a large ancestral cell engulfed a smaller prokaryotic cell, establishing a mutually beneficial relationship where the smaller cell provided energy or photosynthesis and the larger cell provided protection.

What genomic evidence supports the Endosymbiotic Theory for mitochondria and chloroplasts?

They contain their own independent DNA, which is distinct from the nuclear DNA of the host cell.

Do animal cells possess a cell wall?

No, animal cells do not have a cell wall.

Where is the plant cell wall located and what is its primary function regarding osmosis?

It is located outside the cell membrane and keeps cellular pressure under control during osmotic changes.

Why does unicellular yeast share a complex cellular organization with higher organisms like humans?

Because yeast is a eukaryote containing membrane-bound organelles and compartmentalized cellular structures.

What is the hierarchical sequence of structural organization in multicellular organisms depicted here?

Cells $\rightarrow$ Tissues $\rightarrow$ Organ $\rightarrow$ Organ System $\rightarrow$ Organism.

Which cellular components and organelles are labeled in this diagram of a yeast cell?

Periplasm, peroxisome, nucleus, endoplasmic reticulum (ER), secretory vesicles (S), cytosol, Golgi, vacuole, mitochondrion, plasma membrane, and cell wall.

What are four key advantages of cellular compartmentalization and organelles?

1. Creation of unique microenvironments with specific properties (such as pH); 2. Increased concentration of reagents to favor enzymatic reactions; 3. Protection of nuclear DNA; 4. Sequestration of toxic compounds.

What are two disadvantages of cellular compartmentalization?

1. High demand of energy requiring evolutionary adaptation; 2. Increased complexity that makes cellular safeguarding more difficult.

What is a virus in terms of structural composition?

Genetic material (RNA or DNA, single- or double-stranded) encased within a protective protein coating called a capsid.

How does the physical size of a virus compare to that of a bacterium?

Viruses are up to one order of magnitude smaller than bacteria.

What are the four morphological types of viruses?

Helical viruses (e.g., Tobacco Mosaic Virus), polyhedral viruses (e.g., adenoviruses), spherical viruses (e.g., Coronavirus), and complex viruses (e.g., bacteriophages).

What are the two primary classification systems used for viruses?

1. According to shape (helical, polyhedral/icosahedral, complex, spherical); 2. According to genome (single- or double-stranded, positive or negative sense, RNA or DNA).

What is the difference between antigenic drift and antigenic shift in viral evolution?

Antigenic drift refers to small, gradual changes in viral surface proteins caused by mutations over time; antigenic shift refers to a major change when different viral strains recombine genetic material to create a new subtype.

What are two examples of beneficial or symbiotic effects viruses can provide to their hosts?

1. Providing plants with resistance to harsh environmental conditions; 2. Assisting in mammalian placenta formation.

What are three major applications of viruses in biotechnology?

1. Gene Therapy (using adenoviruses or retroviruses as vectors); 2. Vaccine Development; 3. Phage Therapy (using bacteriophages to kill target bacteria).

What three characteristics made SARS-CoV-2 capable of causing a global pandemic?

High transmissibility, asymptomatic transmission, and rapid viral mutation producing new variants.

How are genes, chromosomes, and the genome conceptually explained through the library analogy?

A gene is a single instruction book, a chromosome is a shelf full of books, and the genome is the entire library.

What is the difference between coding genes, non-coding genes, and non-transcribed DNA?

Coding genes encode proteins; non-coding genes are transcribed into functional RNA but not translated into protein; non-transcribed DNA serves structural functions (such as replication or histone binding) or has unknown roles.

Why is a DNA molecule negatively charged?

Because of its phosphate groups, which give the sugar-phosphate backbone a net negative charge and cause it to behave as a mild nucleic acid.

What chemical components form a DNA nucleotide

A phosphate group, a deoxyribose pentose sugar, and a nitrogenous base (A, C, G, or T)

How many hydrogen bonds form between A-T versus G-C base pairs in DNA?

Adenine and Thymine form $2$ hydrogen bonds, while Guanine and Cytosine form $3$ hydrogen bonds.

Why is an A-T base pair chemically weaker than a G-C base pair?

Because $A\text{--}T$ is held together by only $2$ hydrogen bonds, whereas $G\text{--}C$ is held together by $3$ hydrogen bonds.

Why is the principle of base pair complementarity fundamental to modern biotechnology?

It means that if one part of the DNA is missing in one strand, the information is in the complementary bases in the other strand.

In biotechnology, the principle of base pair complementarity allows the recognition of specific DNA fragments and the generation of hybrids between RNA and DNA

What are the three chemical differences between RNA and DNA?

1. RNA contains ribose sugar instead of deoxyribose; 2. RNA contains the base uracil (U) instead of thymine (T); 3. RNA is single-stranded, whereas DNA is double-stranded.

What are the three structural classes of proteins and four general functions they perform?

Classes: fibrous, globular, and transmembrane proteins. Functions: structural support, molecular transport across membranes, enzymatic catalysis, and receptor signaling.

What four steps of enzyme-substrate interaction are illustrated in the Induced Fit Model diagram?

1. Substrate approaches the active site; 2. Enzyme shape changes slightly as the substrate binds; 3. The enzyme-substrate complex forms; 4. Substrate is converted to products, which are released, leaving the enzyme ready for another cycle.

What is Francis Crick's Central Dogma of Molecular Biology (1958)?

Information transfers from nucleic acid to nucleic acid (DNA replication, transcription) or from nucleic acid to protein (translation), but information transfer from protein to protein or from protein to nucleic acid is impossible.

Why can information not travel backwards from a protein to RNA or DNA?

Because of the degeneracy of the genetic code (multiple codons specify the same amino acid) and post-translational modifications, making reverse reconstruction impossible.

What does semi-conservative replication mean in the context of DNA synthesis?

Each newly synthesized double-stranded DNA molecule consists of one conserved parental strand and one newly formed daughter strand.

What enzyme synthesizes the new DNA polymer during replication and what substrates does it assemble?

DNA polymerase, which incorporates free deoxynucleotides (A, C, T, G) using a single strand of DNA as a template.

Where in the genome does DNA replication begin?

At a specific designated sequence known as the origin of replication.

In which direction is an mRNA molecule built during transcription?

In the $5' \rightarrow 3'$ direction.

What is the difference between the sense strand and antisense strand of DNA during transcription?

The antisense strand serves as the template transcribed by RNA polymerase; the sense strand is the non-transcribed strand containing the sequence identical to the resulting mRNA (with T instead of U).

What are the three core stages of transcription?

Initiation - specific nucleotide sequences (Promoters) which indicate RNA polymerase where to begin the RNA synthesis

Elongation - transcription factors unwind the DNA strand to allow the RNA polymerase enzyme to transcribe a single strand of DNA into a single of strand of RNA polymer.

Termination - specific nucleotide sequences (Terminators) that indicate RNA polymerase where to stop the RNA synthesis and detach itself from the DNA

What key regulatory and coding segments compose a basic protein-coding gene?

A promoter, a ribosome binding site (RBS), a coding region (CDS) bounded by Start and Stop codons, and a transcriptional terminator.

What is the primary function of a promoter in a gene?

It serves as the binding site for RNA polymerase and transcription factors to initiate transcription and regulate where, when, and how much mRNA is synthesized.

What is the role of transcriptional terminators located in the 3' UTR?

They provide specific nucleotide sequence signals that instruct RNA polymerase to halt RNA synthesis and dissociate from the DNA strand.

How can genes located on different chromosomes be coordinately regulated in eukaryotes?

By sharing the same regulatory promoter/enhancer DNA sequences that bind identical transcription factor proteins.

What is differential gene expression in multicellular eukaryotes?

The process by which all cells maintain the same complete genome, but specific subsets of genes are turned on or off via cell-specific transcription factors to generate distinct cell types.

How do ribosomes translate mRNA into a functional protein chain?

The ribosome reads the mRNA in triplets (codons), matches each codon to a complementary tRNA anticodon carrying a specific amino acid, and catalyzes peptide bond formation to assemble a polypeptide chain.

What are the universal start codon and three stop codons in mRNA translation?

Start codon: AUG; Stop codons: UAA, UAG, and UGA.

Where in eukaryotic cells does protein glycosylation take place?

Along the secretory pathway inside the endoplasmic reticulum (ER) and the Golgi apparatus.

What is a bacterial plasmid?

A small, circular extrachromosomal DNA molecule that replicates independently of the bacterial chromosome, can exist in multiple copies, and can be horizontally transferred between bacteria.

What is the relationship between genotype, environment, and phenotype illustrated in this slide?

The genotype is the set of genes in an organism, which interacts with the environment to produce the observable phenotype, but only DNA is transmitted to the next generation.

Why does a Himalayan rabbit reared at 20°C develop dark fur on its extremities, while one reared above 30°C remains completely white?

Gene C, which controls fur pigmentation, is active only between $15^\circ\text{C}$ and $25^\circ\text{C}$. Pigmentation appears on ears, nose, and feet at $20^\circ\text{C}$ because those areas lose heat, whereas at temperatures above $30^\circ\text{C}$, Gene C is inactive everywhere.

How does the absence of light affect the phenotype and growth habit of seedling plants?

In the absence of light, chlorophyll expression is not induced, causing plants to appear pale yellow and grow taller and thinner in an effort to reach light.

How do seasonal water fluctuations lead to the formation of annual tree growth rings?

Abundant water in spring stimulates the vascular cambium to produce very large xylem vessels, whereas lower water availability in autumn produces smaller, fewer vessels, creating visible annual rings.

How do plants respond phenotypically when continuously agitated in a wave-like system against gravity?

They continuously attempt to reorient and grow vertically, resulting in thicker cell walls and higher overall biomass.

What is bacterial quorum sensing?

A mechanism by which bacteria produce and sense signaling molecules (auto-inducers) to monitor population density and coordinate group behaviors like virulence and bioluminescence.

What are the sequential stages of bacterial biofilm development shown in this diagram?

Attachment of planktonic microbes to a surface $\rightarrow$ Microcolony formation $\rightarrow$ Maturation and stabilization of biofilm matrix $\rightarrow$ Dispersion of biofilm.

What is the technical distinction between an immunoglobulin (Ig) and an antibody?

Immunoglobulin refers broadly to the entire family of glycoprotein defense proteins produced by B cells, whereas antibody specifically denotes the antigen-binding, immune-active forms.

What are the primary functional characteristics of the 5 antibody classes shown in this chart?

IgG: The most abundant antibody in the blood, providing long-term immunity after infection or vaccination

IgM: The first antibody produced during an initial immune response.

What are four primary effector mechanisms by which antibodies neutralize or eliminate pathogens?

Neutralization - binding to the pathogen to prevent it from interacting with the host cell

Opsonization - binding to pathogens to mark them for destruction by immune cells

Activation of the Complement System - activation of a series of proteins that leads to the bursting of the pathogen

Agglutination - clumping pathogens together

What are the structural regions and chains of an antibody shown in this schematic diagram?

Two identical heavy chains - determines the type of antibody

Two identical light chains - paired with the heavy chain to make 2 binding cites

Variable region - at the tips of the Y-shaped antibody, where the antigen binds. This region varies greatly among antibodies to allow specificity for different antigens.

Constant region - The base of the Y-shape, responsible for interacting with immune cells or other parts of the immune system.

How does a lateral flow pregnancy test function?

The test contains antibodies that specifically bind to hCG in urine.

• Urine flows through the test strip.

• If hCG is present, it binds to the antibodies on the test strip.

• This binding produces a visible line, indicating a positive test.

Why is the control line positioned after the test line on a pregnancy test strip?

To ensure that sufficient liquid volume was applied and that the fluid has migrated completely across the entire test zone.

What is the difference between monoclonal antibodies (mAbs) and polyclonal antibodies?

Monoclonal antibodies originate from a single B-cell clone and recognize one unique epitope; polyclonal antibodies are a mixture from multiple B cells that recognize multiple distinct epitopes on the same antigen.

What is an epitope?

the specific part of an antigen that is recognized and bound by an antibody, a B cell receptor, or a T cell receptor. It is a small molecular structure on the surface of an antigen.

Lock & Key system

What are three mechanisms through which monoclonal antibodies operate in targeted therapies?

1. Neutralizing pathogens by preventing cellular entry

2. Targeting cancer cells to mark them for destruction

3. Blocking inflammatory signals for autoimmune diseases

What three basic functional components are included in a vaccine formulation?

1. Antigen - generates the immune response

2. Preservatives - prevent the vaccine from becoming contaminated once the vial has been opened if it will be used for vaccinating more than one person

3. Stabilizers - prevent chemical reactions from occurring within the vaccine and keep the vaccine components from sticking to the vaccine vial.

What are first, second, and third-generation vaccines?

First-generation: whole virus vaccines (inactivated or live attenuated)

Second-generation: a part of the pathogen

Third-generation: using genetic material like RNA, DNA, and mRNA to instruct body cells and get an immune response.

What is antigenic variation and how does it challenge vaccine design?

A process where viruses frequently alter surface proteins or sugars, enabling them to escape recognition by existing vaccine-induced antibodies.

What three laboratory strategies are used to attenuate live viruses for vaccines?

1. Serial passage in non-human animal cells (virus is passed through another species cell first)

2. Direct genetic modification;

3. Selection of heat-sensitive/cold-adapted strains that replicate poorly at normal human body temperature ($37^\circ\text{C}$).

What are the six sequential stages involved in bringing a vaccine to market?

1. Exploratory stage

2. Proof of concept

3. Testing the vaccine (clinical trials)

4. Manufacturing process

5. Approving the vaccine

6. Distribution.

What is the sequence of IgM and IgG production following vaccination or infection?

IgM antibodies appear first as the initial line of defense, followed later by IgG antibodies, which provide long-term immunological memory.

What is a constitutive promoter and what types of genes does it typically control?

An unregulated promoter that drives continuous, steady gene expression in all tissues; typically controls essential housekeeping genes.

Why are viral promoters frequently employed in biotechnology expression vectors?

Because they are exceptionally strong constitutive promoters capable of driving high levels of transgene transcription.

How does a positive inducible promoter switch from an OFF state to an ON state?

In the OFF state, the activator protein cannot bind the promoter; when an inducer binds the activator, the activator changes conformation, binds the promoter, and initiates transcription.

How does a negative inducible promoter switch from an OFF state to an ON state?

In the OFF state, a repressor protein is bound to the promoter, blocking RNA polymerase; when an inducer binds the repressor, the repressor detaches, allowing transcription to begin.

Why is promoter strength carefully matched to host cells in recombinant protein manufacturing?

It must be strong enough to ensure high product accumulation, but not so excessively strong that it causes metabolic burden or cellular toxicity.

Why can prokaryotic promoters not be used to drive expression in eukaryotic hosts?

Bacterial promoters have simpler consensus elements recognized only by bacterial sigma factors and RNA polymerase, lacking the complex motifs required by eukaryotic transcription factors.

What are the four procedural steps required to produce a transgenic organism?

1. Cloning the gene(s) of interest;

2. Transforming the transgene into the host;

3. Selecting transformed cells

4. Analyzing the modified organism.

What are the two structural segments of a recombinant cloning vector shown in this diagram?

The vector backbone (carrier plasmid DNA) and the insert (the foreign transgene of interest).

What three essential genetic elements must be present within an effective cloning vector?

1. Has to be able to replicate on its own with the transgene and the foreign

   DNA segment that they carry

2. should have a unique site for the insert

3. A selectable marker gene (such as an antibiotic resistance gene).

Why is a selectable marker gene mandatory on cloning vectors?

Because transformation efficiency is far below $100\%$, necessary for the selection of transformed clones AND for the maintenance of the plasmid in the cell (for bacteria)

How does electroporation facilitate the entry of foreign DNA into host cells?

An electrical pulse transiently creates pores in the plasma membrane, enabling negatively charged plasmid DNA to pass into the cytoplasm before the membrane reseals.

What standard thermal parameters define bacterial heat-shock transformation?

Incubation at $42^\circ\text{C}$ for $40\text{ seconds}$.

How is Agrobacterium tumefaciens utilized as a plant transformation vector?

It is a gram-negative soil bacterium with the natural capacity to transfer a portion of its plasmid DNA (T-DNA) into the plant genome, which scientists engineer to deliver transgenes instead of crown gall disease genes.

How does particle bombardment (the gene gun method) introduce DNA into plant tissue?

Microscopic gold or tungsten particles coated with DNA are accelerated at high pressure toward plant tissues, penetrating the cell wall and entering the nucleus.

What are plant protoplasts and how are they generated?

Plant cells that have had their rigid cellulose cell walls enzymatically digested away, leaving spherical, membrane-bound cells capable of taking up DNA via PEG or electroporation.

What two levels of verification are performed when analyzing a newly generated transgenic organism?

Genetic level (confirming transgene presence, completeness, and copy number) and phenotypic level (assessing expected product accumulation, activity levels, and unexpected traits).

Why is the sweet potato recognized as an example of natural transgenesis?

Because sweet potato genomes acquired Agrobacterium T-DNA naturally thousands of years ago during their evolutionary domestication.

What source organism provided the insecticidal gene introduced into Bt corn and cotton?

Bacillus thuringiensis (Bt), a soil bacterium that synthesizes insecticidal crystal proteins.