Dna manipulation - uses enzymes (imitates what cells can do) Restriction endonuclease - able to cleave dna at specific places restriction sites - where nucleases cleave dna. Methylation - stops nucleases from cleaving dna. Type i - makes simple cuts on both dna strands. Type ii - makes staggered cuts where sequences same on both sides (dyad symmetry) ligase - makes phosphodiester bonds to connect hydroxyl/phosphate groups. Also joins okazaki fragments on lagging strands. Creates recombinant molecules from fragments created by nucleases vector systems - used to carry recombinant dna molecule into a cell. Not required by the cell, but can be selected w/ addition of marker. Must have origin of replication, selectable marker (usually for antibiotic resistance) Markers - used to see which cell took in the new dna. Multiple cloning site (mcs) - region in plasmid where dna is inserted inactivation of gene signals plasmid"s acceptance of new dna.