plasmids

**Definition and structure:** Plasmids are small, circular, double-stranded DNA elements found mainly in bacteria. They have their own origin of replication and replicate independently of the bacterial chromosome. They are not required for basic survival but give a selective advantage under certain conditions. Some bacteria (e.g., Borrelia burgdorferi) have linear plasmids. Yeast and other eukaryotes can also carry plasmids.

**Transfer: conjugation (the main plasmid mechanism)** - Requires a sex (F) pilus, encoded by the tra genes on the F (fertility) plasmid. - F+ (donor) x F− (recipient): the F plasmid is transferred, and the recipient becomes F+. The chromosome is usually NOT transferred. - Hfr (high-frequency recombination) cell: the F plasmid is integrated into the chromosome. Conjugation transfers chromosomal DNA, and the recipient rarely becomes F+ because the transfer is usually incomplete. - F' (F prime) cell: the F plasmid excises imprecisely and carries a piece of chromosomal DNA. Transfer of this plasmid gives the recipient bacterial genes. - Plasmids can also move by transformation (uptake of naked DNA). Transduction occasionally carries plasmids as well, but this is less commonly tested.

**Major types and clinical examples** - **R (resistance) plasmids:** carry genes for antibiotic resistance, such as β-lactamases, aminoglycoside-modifying enzymes, and efflux pumps. They can carry multiple resistance genes at once, so multidrug resistance can transfer in one conjugation event. Examples include ESBL and carbapenemase genes (e.g., KPC) in Enterobacterales, and vanA in VRE. - **Virulence plasmids** encode toxins and other virulence factors: - Bacillus anthracis: pXO1 (toxin: protective antigen, edema factor, lethal factor) and pXO2 (capsule) - Clostridium tetani: tetanospasmin - ETEC: heat-labile (LT) and heat-stable (ST) toxins - Yersinia: Yops - Shigella and EIEC: invasion plasmid - **Col plasmids** encode bacteriocins, which kill competing bacteria. - **Metabolic or degradative plasmids** encode enzymes for degrading unusual substrates (e.g., toluene in Pseudomonas).

**Comparison with other mobile elements** - **Transposons ("jumping genes")** move from one DNA site to another. They can integrate into plasmids and carry resistance genes between plasmids and chromosomes. - **Bacteriophages** carry the toxin genes of the lysogenic conversion group. The mnemonic is "ABCDEFG": Shiga-like toxin (EHEC), Botulinum toxin (C. botulinum), Cholera toxin, Diphtheria toxin, Erythrogenic toxin (S. pyogenes), and others. These are phage-encoded, not plasmid-encoded. - Note the bacterial toxins that are plasmid-encoded: tetanus toxin, ETEC LT/ST, and the B. anthracis toxins.

**Biotechnology significance:** Plasmids are the workhorse vectors for recombinant DNA technology. - A cloning vector contains an origin of replication, a selectable marker (usually antibiotic resistance), and a multiple cloning site (MCS) with restriction enzyme sites. - A foreign gene is inserted using restriction enzymes and DNA ligase to make recombinant DNA. The plasmid is then transformed into bacteria and selected on antibiotic plates. - Blue-white screening (lacZ) identifies bacteria with inserts. White colonies have the insert, which disrupts lacZ. - Used to produce recombinant insulin, growth hormone, and hepatitis B vaccine (in yeast). Also used for DNA vaccines and some gene therapy vectors.

**High-yield takeaways** 1. Plasmids carry nonessential genes (resistance and virulence) and replicate independently. 2. Conjugation uses the F pilus. The F+ x F− cross transfers the plasmid only, and the Hfr cross transfers chromosomal genes. 3. Plasmid-mediated resistance spreads horizontally, so one transfer can confer resistance to many drugs. 4. Plasmids are the standard cloning vectors in molecular biology.