Biotechnology and Genetic Engineering: Key Terms and Glossary

The United States decision that a human-altered bacterium could be patent-eligible. It does not authorise release, and other countries do not copy the holding.

The host locus activated by enhancer insertion from those early SCID vectors. The lesson is that the insertion site is part of the pharmacology, not that gene therapy as such cannot work.

The United States decision that an isolated natural gene sequence is not patent-eligible, whereas cDNA can be. Patent practice elsewhere draws its own line between a genomic sequence and a cDNA.

A marker gene used so that rare transformed plant cells can be recovered. Assessments have judged dietary risk from its protein low; the argument that remains is about a drug-resistance gene in the food chain.

Virulence genes that stay on the Ti plasmid outside the transferred DNA, nick and coat the T-strand, and deliver it, so a helper plasmid can supply them in trans.

Adeno-associated virus used as a gene-therapy vector. Its genome stays mostly episomal, the cargo is small, the DNA is lost in rapidly dividing cells, and pre-existing immunity is common.

Use of a chemical analogue to enrich mutants that ignore feedback on a biosynthetic enzyme, or that merely alter transport. Only a product assay tells those outcomes apart.

The antimalarial precursor produced in yeast from farnesyl pyrophosphate by a plant-derived route. It is not artemisinin, chemistry finishes the drug, and a laboratory titre is not a supply chain.

The February 1975 meeting that considered lifting the pause by matching experiments to physical and biological containment. It was not a ban, not a regulator, and not a statute still in force.

A bacterial artificial chromosome maintained at low copy from an F origin, suited to inserts on the order of a hundred to a few hundred kilobases and cleaner than a yeast artificial chromosome.

An insect-cell virus used for expression by placing the foreign gene under a very late, very strong promoter. The yield is high and the infection kills the culture.

A nickase fused to a deaminase, changing one class of base inside a short window without a clean double-strand break. Other editable bases in that window can change as well.

A culture in which the nutrients are present at the start and the volume is fixed, so growth follows a batch curve and there is no steady state.

The 1974 request that scientists voluntarily defer plasmids carrying new resistance or toxin genes, and animal-virus DNA joined to vectors that replicate in bacteria. It was not a statute.

A two-plasmid arrangement in which *vir* functions sit on a helper and the border sequence, with the genes to be transferred, sits on a shuttle plasmid.

Use of a host and a vector that are unlikely to thrive outside the laboratory.

The use of organisms to degrade some organic pollutants when the pathway and the right electron acceptor are both present, or to precipitate or reduce metals whose atoms remain. It is an outline, not a site treatment.

A containment category matched to the hazard of a procedure. The same species can be handled at more than one level, and the level is not a bench manual.

A recognition event, by a protein, riboswitch, enzyme, or antibody, converted into fluorescence, current, or growth. If it also sees an upstream intermediate, selection will accumulate that intermediate.

A colour test in which an insert in the polylinker usually stops α-complementation and leaves a white colony. A white colony reports that the α fragment was interrupted; the identity of the insert is a separate claim.

A flush end that can be joined to any other blunt end, with less efficiency and with no overhang to choose the partner.

The 2000 agreement on transboundary movement of living modified organisms, written in precautionary language. It is not a single national statute and not a rulebook for an insulin factory.

A Cas relative which, in the notes' account of Cas12a, uses a T-rich PAM, cuts staggered ends, and can process its own guide array.

An RNA-guided nuclease of bacterial adaptive immunity, programmable by a guide and still dependent on a PAM and on how the cell repairs the break.

A DNA copy of RNA, usually of mRNA, already spliced and often incomplete at the 5′ end, with abundance following that sample rather than the chromosomal gene.

A clone collection of spliced mRNA from one sample. Abundant transcripts dominate, promoters are absent, and a gene silent in that sample is missing.

A continuous culture in which inflow equals outflow, so the pump sets the growth rate. A faster mutant that makes less product is enriched, not thrown away at the end as in a batch.

An antibody that keeps mouse variable domains and replaces the constant domains with human ones, so less of the mouse sequence remains than in the original hybridoma protein.

An engineered T-cell receptor that joins an antibody-binding domain to T-cell signalling domains, so the cell kills targets that display the chosen antigen. Loss of normal cells that share the antigen is on-target, not an off-target accident.

A mammalian host cell, the usual choice for licensed antibodies because it adds complex N-glycans, which are still not identical to human ones.

The count of clones expected to give a chosen probability of covering a genome, from the insert length as a fraction of that genome.

The deliberate use of an organism, or of a part of one, to make a product, where any genetic change is whichever variant selection happened to enrich and the sequence need not be known.

The 1973 demonstration that plasmid DNA cut and rejoined could be transformed into *Escherichia coli*, replicate, and express a resistance the host plasmid had not carried.

Overhangs that can base-pair even when they were cut by different enzymes, sometimes so that the hybrid junction matches neither parent site.

Removal of a floxed allele only in a chosen tissue or at a chosen time, so the result is not limited to whatever a constitutive null did to the embryo.

How many plasmid copies the origin maintains. A high number yields more DNA and a greater burden, and is a poor place for a toxic product or a huge insert.

A plasmid carrying phage cos sites so that it can be packaged in a phage particle and then replicate as a plasmid, used for inserts of a few tens of kilobases.

The fraction of chains that successfully add the next nucleotide in one synthetic cycle. A small loss compounds with length, so a long chain made in one piece is rich in deletions.

A glyphosate-insensitive EPSPS, often from *Agrobacterium*, expressed so that a crop survives a herbicide that stops the plant's own enzyme.

The cycle at which a qPCR fluorescence signal crosses a chosen threshold. An earlier cycle means more starting template only if the assay is specific and the efficiencies match.

Formation of ethanol by bakers' yeast from excess glucose even when air is present, the overflow fed-batch culture is arranged to avoid.

A recombinase from phage P1 that rearranges DNA only at lox sites already present. The DNA must already carry those sites.

Repression of transcription by a dead Cas protein, optionally fused to a repressor domain. The effect lasts while the protein is present and does not rewrite the DNA.

An insecticidal protein from *Bacillus thuringiensis*, activated in an alkaline insect midgut, where it binds a specific receptor and forms a pore. The spectrum is narrow, and field resistance has evolved where the genetic assumptions failed.

Cas9 with both nuclease sites inactivated, so it binds a target and does not cut. With a fused regulator it can change transcription while the protein is present, and it is not a heritable allele.

An insertion, codon, deletion, or pathway that was chosen on purpose, with the rest of the genome supposed to stay put and still needing to be checked.

The volume fraction replaced per unit time in a chemostat. At steady state it equals the specific growth rate.

A cycle of diversification, expression, and selection or screening, repeated so that the enzyme which emerges is the one the assay rewarded, which may not be the property that was wanted in use.

An enzyme that seals a nick by joining a 3′ hydroxyl to a 5′ phosphate. The T4 enzyme is the cloning ligase because it can seal blunt ends as well as sticky ones.

Recombination among related sequences to generate the diversity for a round of directed evolution. The idea is named here, not a laboratory procedure.

Research that would make a pathogen more transmissible, more virulent, broader in host range, harder to treat or to detect, or possible to reconstitute, or that would provide the design for someone else to do so. It is not a synonym for recombinant DNA, and these notes do not describe how such work is done.

The question of who can obtain a product after it can be made. Access to insulin, to a scarce gene therapy, and to a nutritional crop fail in different ways, and counting the gap is not an argument for skipping regulation.

Genetic modification of cells outside the body, so a failed lot can be discarded and the vector does not have to find those cells inside the patient.

Who meets a hazard, by which route, and how often. Risk is the combination of hazard and exposure, so a contained organism and a released one are not the same risk.

An enzyme from an organism that lives at high temperature, high pH, or another extreme. The niche suggests a stability and does not prove expression yield or performance in a process broth.

A clotting factor once prepared from pooled human plasma, which transmitted blood-borne viruses to many recipients, and now made from a recombinant gene so that particular risk is removed.

A culture to which feed is added and whose volume rises, growth being limited by the feed. It is how excess substrate, and overflows such as the Crabtree effect, are avoided.

An allele flanked by lox sites so that Cre can delete it only where, or after, the recombinase is expressed.

The sharing of control along a pathway, so that overexpressing one enzyme which had been called rate-limiting often changes the flux very little.

The nuclease domain fused to zinc-finger and TALE arrays. It cuts as a dimer, so one monomer bound at a partial site does not cut.

A single polypeptide that joins two functions, such as a receptor domain and an antibody Fc region. The partner can mask a signal, and the result is a new protein.

Movement of a gene into another sexually compatible population. Probability depends on compatibility, distance, and flowering; pollen moves an allele whatever legal label the allele carries.

Delivery of DNA or protein on particles shot into a cell, including into chloroplasts, which T-DNA does not target. The idea is named here, not a protocol.

A clone collection that represents total DNA, including introns and promoters, sized so that the insert fraction makes coverage of the genome probable.

An edit in a gamete or an embryo, inherited by descendants who cannot consent. Efficiency does not close that question, and an accidental gonadal exposure can make a somatic plan germline.

Heritable editing of a human gamete or embryo. A professional norm held that clinical use should wait for safety, a real unmet need, and broad social agreement, and a working method is not permission to cross that line.

Rice engineered so that the endosperm, which does not normally make carotenoid, accumulates β-carotene. It is a nutritional trait, not a pesticide and not a capsule of vitamin A, and approval and adoption differ by jurisdiction rather than standing as one current statute.

The harm an agent or a procedure can cause, considered separately from who meets it and how often.

Repair that copies a homologous donor, so a designed allele can be introduced, mostly in dividing cells and usually less often than end joining in mammalian cells.

A biosynthetic route moved into a host that did not possess it, and that still has to compete with the host's own metabolism and with toxic intermediates.

The plan, for an insecticidal crop, that the dose kills heterozygotes so resistance is functionally recessive, while a non-expressing refuge supplies susceptible mates. It fails if the dose is low for that pest, resistance is not recessive, or the refuge is absent.

A short run of histidines, commonly six, that binds an immobilised metal and can be displaced by a competitor. Elution shows that the handle is present, not that the protein is active.

Grafting mouse complementarity-determining regions onto a human antibody framework. Framework residues are often restored because grafting alone can drop affinity.

Detection of colony or plaque DNA by a labelled nucleic acid. A hit can be a fragment, an intron, or a mutated copy, depending on the stringency.

A fusion of one antibody-secreting B cell with a myeloma cell, immortalising a single specificity.

The production phase in that classical schedule, following the growth phase, and a simplification of how secondary metabolism is actually regulated.

An intracellular aggregate formed when synthesis outruns folding, especially for disulphide proteins made in the reducing bacterial cytoplasm.

Agreement, without coercion and in understandable language, covering purpose, procedure, risks, alternatives, and the right to withdraw. Vector risks are part of what must be said.

Damage or activation of a host gene by where a vector integrates. Early gamma-retroviral treatment of X-linked SCID restored immunity and, in several children, caused T-cell leukaemia by enhancer insertion near *LMO2*.

A restriction enzyme that recognises the same sequence as another.

An integrating viral vector with a larger cargo than AAV, so daughter cells inherit both the added gene and whatever insertion was made.

Single-molecule reads long enough to show rearrangements and whether a plasmid backbone is the one that was drawn.

The 34 bp site Cre recognises, inverted repeats around a directional spacer. Same orientation excises the interval; opposite orientation inverts it.

The joining of DNA from different sources and its return to a cell as a molecule that can replicate, so a specified fragment can sit with a known origin and a known marker.

That voluntary pause, not a claim that every recombinant molecule was dangerous and not a law.

The presence of more than one genotype in a treated founder. Breeding and genotyping, not the act of injection, are what produce a clean line.

A short run of unique restriction sites at which the circle can be opened without destroying the origin or the marker.

End joining that works in most cells, including non-dividing ones, and often leaves a small, unpredictable indel. It can knock a gene out of frame and cannot write one specified codon.

A Cas9 variant in which one of the two nuclease sites is mutated, so the enzyme cuts only one strand. Paired nickases demand two binding events.

The same question asked of RNA: whether a transcript of the expected size is present.

A cut at a site other than the one intended, more readily tolerated away from the PAM-proximal seed. A predicted list is not a clearance; the cuts have to be measured.

The sequence without which a vector is not copied as the cell divides, so the DNA is diluted away.

Movement of oxygen from the gas phase into the liquid, which limits a dense aerobic culture because the dissolved stock is small and is consumed quickly. It is a constraint, not an operating instruction.

A short motif that a Cas nuclease requires beside the target sequence, which is why a CRISPR array can store spacers without cutting its own genome. It is not a guide sequence, and none is given here.

A temporary right, in the country that grants it, to exclude others in exchange for disclosure. It is not ownership of a wild species and not permission to release an organism or to sell a medicine before a regulator agrees.

Resistance conferred by a sequence from the pathogen itself, as in coat-protein transgenic papaya, acting largely through RNA silencing. Recovery of that industry does not generalise to every virus or every crop.

Repeated copying of whatever DNA actually lies between two primers, each product becoming a template, so the gain is exponential and a contaminant can become the whole signal. Amplified does not mean faithful, and no primer sequence is specified here.

A selection in which a peptide or protein is shown on a phage and the gene is kept if the product binds a surface. Binding is not serum stability and not catalysis unless the scheme was built to demand them.

Production of a therapeutic protein in the milk or another product of a transgenic animal, a choice of host when amount or modification suits a mammal, not a new kind of genetics.

Chemical assembly of DNA on a solid support, one nucleotide per cycle and in the opposite direction from a polymerase. It writes a specified short strand and does not judge whether that strand should be made.

The barriers of the laboratory, as distinct from the biology of the host and the vector.

A drawing of origin, marker, and expression cassette that is tested by whether the predicted cuts appear and by reading the cassette in transcription order.

Enrichment for a targeted change in embryonic stem cells by selecting for a cassette inside the homology and against a marker that survives only if insertion was random.

The stance that a threat of serious or irreversible harm is not postponed merely because certainty is incomplete, and, in stronger forms, that the proposer must show sufficient safety first.

A product tied to growth, such as an amino acid, a central organic acid, or ethanol, and normally regulated so that the cell does not make a large surplus.

A nickase fused to a reverse transcriptase, copying a new sequence from an extended guide after a nick, so a small change need not start from a double-strand break.

The single-chain insulin precursor that folds with its connecting peptide in place. The peptide is removed afterwards, avoiding the poor yield of correct disulphides when the A and B chains are joined separately.

Introduction of DNA into a fertilised egg's pronucleus, integrating at random and often as a multi-copy array, so each founder differs and several are required before a phenotype is blamed on the transgene.

A change at a residue a structure suggests. A substitution that kills activity falsifies that particular guess and does not, by itself, explain why.

A molecule made by joining DNA from different sources so that it can be propagated as one replicon.

A bacterial defence that cuts incoming DNA at particular sequences and methylates the same sequences on the cell's own genome.

A polymerase that copies RNA into DNA, the step required before PCR or cloning can use an RNA template.

A nuclease delivered already assembled with its guide, so the activity disappears as the protein is degraded rather than persisting from an expressed gene.

A comparison of estimated harms and benefits, including harm from leaving things as they are. It genuinely disagrees with precaution about who must prove the case and about whether delay itself counts as harm.

Chain termination with dideoxynucleotides, reading on the order of several hundred to about a thousand bases from one clean template, enough for a junction or a short insert and not, by itself, for a long unchecked middle.

A later question put to colonies that have already grown, asking which of them carries the intended molecule or activity.

A product, many antibiotics among them, that is not required for growth in the laboratory and classically appears as growth slows, so forcing growth can lower the titre.

A gene, often a drug resistance or a nutritional complement, that allows only cells which received the vector to form a colony. It does not prove that an insert is present.

An organism that passed a test. The bases that differ, and which of them cause the phenotype, need not have been specified in advance.

A condition that decides which cells form a colony, such as an antibiotic matching the vector's marker. Empty vector and wrong insert both survive it.

The bacterial ribosome-binding site placed a few nucleotides upstream of the start codon. A eukaryotic message needs a Kozak context instead.

Massively parallel reads of roughly a hundred to a few hundred bases, whose depth can catch rare variants in a plasmid population.

A vector with a second origin, so the same molecule can replicate in more than one kind of host.

An edit in body cells that ends with that person, provided it has not reached the germline. The person can usually be asked; off-target alleles and mosaicism remain.

Placement of a somatic nucleus into an enucleated oocyte. Dolly, reported from an adult mammary cell, showed that cloning copies a genome; it is not genome editing, even if an edit and a transfer are later stacked.

A membrane hybridisation that asks whether a DNA sequence is present and on which fragment. It does not show a safe organism or a balanced pathway.

Cutting at sites related to the canonical site when the reaction conditions are poor, which creates fragments the map did not predict.

An overhang left by a staggered cut, able to base-pair with a matching overhang and so make ligation both efficient and selective.

How similar a sequence must be before a hybridisation probe remains bound. Lower stringency buys distant homologues and also false positives.

The DNA, bounded by short repeats, that *Agrobacterium* transfers as a single strand into the plant nucleus, where it integrates largely at random. Only that strand is delivered; the bacterial cell stays outside the plant cytoplasm.

A nuclease guided by TALE repeats, more nearly one repeat per base than a zinc finger, still fused to a FokI dimer and still dependent on cellular repair.

The heat-stable DNA polymerase from *Thermus aquaticus* that survives the denaturation step and so made PCR routine. It lacks proofreading.

Treating an early trial as if it were care offered because it will help the subject, a confusion that serious illness makes more likely.

The *Agrobacterium* plasmid that carries both the DNA to be transferred into a plant and the virulence functions that send it.

A restriction enzyme that cuts within or beside a short, defined site and usually needs only magnesium, which is why this class became the cloning cutter.

The virulence protein that nicks a T-DNA border and remains covalently attached at the 5′ end of the transferred strand.

Loss of the culture when the dilution rate exceeds the organism's maximum specific growth rate and cells leave faster than they divide.

A probe of protein, usually with an antibody, asking whether a polypeptide of the expected size is there. An abundant mRNA can exist without that band.

A yeast artificial chromosome that can carry very long DNA and that often joins two unrelated fragments, which is why it was a poor final map unit.

A pair of zinc-finger DNA-binding arrays, each fused to FokI, aimed at a chosen site. The idea is a protein-guided break; each new target was a new protein, and the cell's repair makes the edit.

Restoration of β-galactosidase activity when a plasmid-borne α fragment supplies what a host *lacZ* deletion lacks, the interaction a blue–white test interrupts.

The provitamin A accumulated in Golden Rice grain. It is not retinol, and conversion in the body is not the same thing as swallowing preformed vitamin A.