1. Plasmid biology: plasmids are extrachromosomal elements that can replicate independently of the host cell chromosome (generally do not contain any essential genes for the host cell's survival)

plasmids are distinguished from one another in several ways, noted below.

-size: from 1 to >100 kb

-copy number: from 1 to >30/cell (size usually inversely proportional to copy #).

-host range: the species/genera of bacteria in which a plasmid can stably replicate.

-incompatibilty group: that is, can two plasmids be stably maintained in same cell. If 2 different plasmids can co-exist in the same cell, they are in different incompatibility groups.

-other traits: includes whether a plasmid is conjugal, carries antibiotic resistance genes, confers phage sensitivity, encodes bacteriocin genes, etc.

2. Conjugation: gene transfer from donor to recipient requiring cell to cell contact which is mediated by the sex-pilus. Amount of genetic information transferred can be significantly larger than in transformation or trandsduction.

During conjugation, generally a plasmid transferred to recipient. (Can also mediate transfer of other non-conjugal plasmids that may reside in same cell).

Conjugation mechanism for F plasmid (and probably other Gram neg. plasmids).

• initial contact mediated by conjugal pilus.
• establishment of mating pairs after retraction of pilus.
• conjugal DNA processing&emdash;plasmid nicked in site- and strand-specific way at oriT (origin of transfer). Single strand of plasmid DNA transferred in 5' to 3' direction to recipient cell, followed by re-ligation of nicked DNA.
• replacement DNA strand synthesis occurs in both the donor (leading strand) and the recipient (lagging strand) cell, concurrently with DNA transfer.
• after mating pair separate, both the donor and the recipient contain the conjugal plasmid.

1. Plasmids can integrate into the chromosome of host cell via transposable elements (either insertion sequences-IS- or transposons-Tn). This can happen at low but detectable frequency. If an F plasmid remains in the E. coli chromosome, the cell is called an Hfr.

If the F plasmid inaccurately excises from the chromosome after formation of an Hfr, it can take a portion of the chromosome with it, which then becomes part of the plasmid itself. This form of the F plasmid is called an F' (F prime).

2. Transfer of chromosomal DNA from a donor to a recipient can occur by transfer mediated by an Hfr (where the chromosomal DNA transferred dependent on the site of the F plasmid insertion into the chromosome) or by an F' (where only a restricted amount of chromosome carried on the plasmid is transferred).

Restriction-Modification systems:

Bacteria can protect themselves from foreign DNA using restriction-modification systems. If incoming DNA introduced into the cytoplasm of the cell via transformation, or conjugation does not have a characteristic pattern of post-replication modification (usually methylation at either A or C bases within a particular DNA sequence), the incoming DNA is susceptible to endonucleases in the cell.

The nucleases that carry out the degradation of foreign DNA are generically called restriction enzymes (frequently, the DNA cutting or restriction enzyme is separate from the DNA methylation or modification enzyme). Restriction enzymes that cleave unmodified DNA at predictable sequences have been useful for the constuction of engineered plasmids for biotechnology.