Ch. 17

• restriction enzymes


  identify specific DNA sequences (recognition sites) and cleave four to eight base pairs into fragments
• reverse transcriptase 


  processed mRNA can be used as a template for complementary DNA synthesis, which can then be cloned without the need for RNA processing; isolated from retroviruses
• gel electrophoresis


  DNA fragments are placed at the negative end of an electrical field and they migrate toward the positive pole; migration rate is determined by fragment size and gel density
• southern blotting


  distinguishes fragments of DNA; DNA molecules are separated through electrophoresis, transferred to a nylon membrane and bathed in a radioactive solution containing a probe (fragment of DNA complementary to the DNA of interest)
• polymerase chain reaction


  enables the rapid synthesis of billions of copies of a specific DNA fragment; process consists of denaturation, annealing, and extension and requries primers, nucleotides, polymerase, and template strands
• taq polymerase


  commonly used in PCR due to its ability to function in high temperatures
• cloning vectors


  replicates host organism for genes to be cloned; ex. plasmids, phages, cosmids, and artificial chromosomes
• cosmids


  maintain fragments of cloned DNA that are too large for plasmids or phages; consist of a cos site from a phage and selectable marker, ori, and MCS from a plasmid
• artificial chromosomes


  used when constructing a genomic library or sequencing an organism's entire genome; ex. bacterial and yeast
• selectable marker


  a gene in a plasmid that encodes a protein that is needed for the cell to survive under certain selective conditions
• muticloning site (MCS)


  many unique restriction sites clustered in a single region so a fragment of DNA can be cloned into a plasmid
• genomic library


  used to clone a gene based on function when the actual sequence is unknown; the organisms genome is cut into fragments, cloned into vectors, and carried by different microorganisms in order to isolate the target gene sequence
• heterologous gene


  when a gene from one organism is cloned into another; many cannot be expressed without further modification
• expression vectors


  contain protomers that result in high-level transcription of heterologous genes; often include regulatory regions (inducers) for health of host cell
• polyhistidine tagging


  adding a series of histidine amino acid residues to either the N or C terminus of the protein; his-tagged proteins bind to a resin and are then seperated from other cellular constituents