Chapter 8: Analyzing Cells, Molecules, and Systems - Vocabulary Flashcards

A set of 50 vocabulary flashcards covering key methods, concepts, and terminology from Chapter 8 on analyzing cells, proteins, nucleic acids, and genetics.

Primary culture

Cell cultures prepared directly from the tissue of an organism, which divide for a limited number of times in culture.

Replicative cell senescence

The protective mechanism in which normal primary cell cultures stop dividing after a limited number of cell divisions.

Immortalized cell lines

Cell lines that are capable of dividing indefinitely in culture, often derived from tumors or transformed by expressing telomerase or tumor-inducing viruses.

Culture shock

A process by which cultured cells stop dividing despite telomerase activity due to protective mechanisms activated by excessive mitogenic stimulation.

Fluorescence-activated cell sorter (FACS)

An instrument that monitors individual cells for fluorescence as they pass through a laser beam, assigns electrical charges to single-cell droplets, and deflects them into collection tubes using an electric field.

Hybridoma cell line

A hybrid cell line produced by fusing a B lymphocyte from an inoculated mouse with a transformed B lymphocyte line to continuously produce specific monoclonal antibodies.

Monoclonal antibodies

Identical antibodies produced by a single clone of hybridoma cells that specifically bind to a single antigenic site.

Differential centrifugation

A procedure that separates cellular components and organelles in cell homogenates based on size and density through a series of increasing centrifugal speeds.

Velocity sedimentation

A separation method where cell components are layered onto a shallow sucrose gradient ($5\text{ to }20|\times 10^{-2}$) and separated by size and shape during centrifugation.

Equilibrium sedimentation

A separation method where cellular components migrate through a steep sucrose or cesium chloride gradient ($20\text{ to }70|\times 10^{-2}$) until reaching a position corresponding to their buoyant density.

Column chromatography

A purification technique in which proteins in a mixture are separated based on chemical properties such as charge, hydrophobicity, size, or binding affinity as they pass through a porous matrix.

High-performance liquid chromatography (HPLC)

An advanced chromatographic method using high pressure and tightly packed small matrix beads to separate proteins with high resolution.

Ion-exchange chromatography

A type of column chromatography that separates proteins based on surface net charge using charged matrix beads.

Gel-filtration chromatography

A type of column chromatography that separates proteins based on size using porous matrix beads that retard small molecules while allowing large molecules to elute unretarded.

Affinity chromatography

A protein purification method using matrix beads covalently linked to a specific substrate, ligand, or antibody to selectively bind target proteins.

Immunoprecipitation

A purification method using specific antibodies bound to heavy beads to pull specific target proteins out of a complex cell homogenate mixture.

Tandem affinity purification tagging (TAP)

A protein isolation strategy employing two sequential affinity tags separated by a specific protease cleavage site to gently isolate native protein complexes.

SDS-polyacrylamide gel electrophoresis (SDS-PAGE)

An analytical technique used to separate denatured polypeptide chains according to their molecular mass using an electric current across a polyacrylamide gel matrix.

Sodium dodecyl sulfate (SDS)

An anionic detergent used in protein gel electrophoresis to denature proteins and coat them with uniform negative charges proportional to their molecular weight.

$\beta$-mercaptoethanol

A reducing agent added to protein samples in SDS-PAGE to break disulfide ($\text{S-S}$) covalent bonds and dissociate multi-subunit complexes into individual polypeptide chains.

Isoelectric point (pI)

The specific pH value at which a protein carries no net electrical charge and therefore stops migrating in an electric field.

Two-dimensional gel electrophoresis

A high-resolution separation technique combining isoelectric focusing (separation by pI) in the first dimension and SDS-PAGE (separation by molecular weight) in the second dimension.

Western blot

An analytical technique where gel-separated proteins are transferred to a membrane and identified using specific antibodies.

Tandem mass spectrometry (MS/MS)

A mass spectrometry technique where specific precursor peptide ions are selected in a first stage, fragmented with inert gas, and analyzed in a second stage to determine amino acid sequences.

X-ray crystallography

A method used to determine the three-dimensional structural arrangement of atoms in a protein by analyzing the diffraction pattern of X-rays directed through a protein crystal.

Restriction nucleases

Bacterial enzymes that cut double-stranded DNA at specific nucleotide recognition sequences, producing blunt or staggered sticky ends.

Agarose gel electrophoresis

A technique used to separate DNA or RNA fragments by size as they migrate through an agarose matrix toward a positive electrode.

DNA hybridization

The specific base-pairing process where a labeled single-stranded nucleic acid probe binds to a complementary DNA target sequence.

Polymerase chain reaction (PCR)

An in vitro method used to rapidly amplify specific target DNA sequences through repeated cycles of denaturation, primer annealing, and DNA polymerase synthesis.

Short tandem repeat (STR) profiling

A PCR-based DNA profiling technique that analyzes length variations at specific short repeated sequence loci to distinguish between individuals.

Plasmid vector

A small circular double-stranded DNA molecule derived from bacteria used to carry and replicate inserted foreign DNA fragments inside host cells.

DNA ligase

An enzyme that completes recombinant DNA construction by catalyzing the covalent linkage of the sugar-phosphate backbones of complementary DNA fragments.

Genomic library

A comprehensive collection of cloned DNA fragments representing the complete genomic DNA sequence of an organism inside plasmid vectors or bacteria.

Complementary DNA (cDNA) library

A collection of cloned DNA molecules synthesized from cellular mRNA using reverse transcriptase, representing only expressed protein-coding genes.

Sanger sequencing

A classic DNA sequencing method that incorporates chain-terminating dideoxyribonucleoside triphosphates (ddNTPs) lacking a 3'-OH group to produce labeled fragments of varying lengths.

Next-generation sequencing

A high-throughput DNA sequencing method (such as Illumina) that incorporates reversible fluorescent terminator nucleotides and records fluorescence emission after each cycle.

Open reading frame (ORF)

A continuous stretch of DNA nucleotides containing a start codon and lacking stop codons, representing a potential protein-coding gene sequence.

Forward genetic screen

An experimental strategy that begins with random mutagenesis to discover organisms exhibiting specific altered phenotypes, followed by identification of the affected gene.

Reverse genetics

An experimental strategy that starts with a known gene sequence, selectively mutates or inactivates it, and examines the resulting phenotypical outcome to deduce gene function.

Point mutation

A genetic mutation that maps to a single site in the genome, corresponding to a single nucleotide pair or a very small part of a gene.

Conditional mutation

A mutation that produces a mutant phenotypic effect only under restrictive conditions (such as high temperature) while exhibiting a wild-type phenotype under permissive conditions.

Loss-of-function mutation

A mutation that reduces or completely abolishes the normal activity of a gene, typically exhibiting recessive inheritance.

Gain-of-function mutation

A mutation that increases the activity of a gene or causes it to become active in inappropriate contexts, typically exhibiting dominant inheritance.

Dominant-negative mutation

A dominant-acting mutation in which the altered mutant gene product interferes with the function of the normal wild-type gene product, causing a loss-of-function phenotype in heterozygotes.

Suppressor mutation

A secondary mutation that suppresses or negates the phenotypic effect of a primary mutation, restoring a normal wild-type phenotype in double mutants.

Complementation test

A genetic procedure in which two homozygous recessive mutant strains are crossed to determine whether their mutations reside in the same gene or in two different genes.

Single nucleotide polymorphism (SNP)

A single nucleotide base-pair variation at a specific genomic location that occurs commonly among individuals within a population.

Cre-loxP system

A site-specific recombination technology used to selectively delete or rearrange target genes flanked by loxP sites in specific cell types or tissues.

CRISPR-Cas9

A targeted genome-editing technique that uses a guide RNA to direct the Cas9 endonuclease to make precise double-strand breaks at specific target genomic sites.

Chromatin immunoprecipitation (ChIP)

A genome-wide method used to map specific DNA sequences bound by transcription regulators in living cells using formaldehyde cross-linking, antibody precipitation, and DNA sequencing.