Gene Expression & Biotechnology

Feedback Mechanisms

  • Inducible: Lac operon (negative control)
    • regulatory gene codes for the active repressor protein
    • repressor attaches to the operator & blocks RNA polymerase from transcribing genes that digest lactose
    • when lactose is present, it binds to the repressor and changes its shape so it can’t attach to the promoter
    • RNA polymerase can then transcribe genes for lactase to digest the lactose
  • Repressible: Trp operon (negative control)
    • usually on
    • regulatory gene codes for inactive repressor protein
    • when tryptophan is present, it acts as a corepressor and binds to the repressor
    • the corepressor changes the repressor’s shape to attach to the promoter → blocks transcription of trp synthesis genes
  • Positive Gene Control
    • catabolic activator protein (CAP):: activates transcription
    • when no glucose present, CAP binds with cAMP and attaches to promoter and accelerates transcription, cAMP=activator
    • when high glucose levels present, CAP detaches

Transcriptional Controls

  • Acetylation:: loosens chromatin, promotes initiation of transcription
  • Methylation:: condenses chromatin, less transcription
  • epigenetic inheritance:: inheriting traits because of factors outside of nucleotide sequence
  • enhancers & activators increase transcription
  • microRNA fragments:: bind to complementary mRNA and silence it by preventing translation

Genes & Expression

  • chromatin:: consists of DNA, forms chromosomes
  • nucleosome:: structural unit of chromosomes, consists of DNA coiled around histones
  • heterochromatin:: very tightly packed DNA, so it has less transcription
  • totipotency:: ability of a cell to divide & produce multiple differentiated cells in an organism
  • nuclear transplantation:: cloning, entire DNA of an organism is used to produce another organism that is genetically identical
  • transposon: repetitive DNA sequence that can move locations, introduces new combinations of DNA → genetic diversity
  • gene duplication:: duplication of DNA that contains a gene, type of chromosomal mutation
  • being heterozygous is more advantageous than homozygous because recessive traits that carry disease aren’t expressed (ex. sickle cell)
  • HOX genes:: control body plans/where parts go in an organism (ex. head on top, legs on bottom, etc)
  • operon:: cluster of genes transcribed together → mRNA → proteins, typically in prokaryotes

Restriction Enzymes

  • “molecular scissors”
  • cut DNA at palindromic restriction sites, sticky or blunt cuts
  • defense against bacteriophages in nature
  • recombinant DNA:: sticky cut DNA fragments can be recombined in different orders, sealed by ligase

Genetic Engineering

  • vectors:: desired genes stored and delivered with viruses, bacteria, etc
  • electrophoresis
    • DNA digested/cut by restriction enzymes is placed into agarose gel exposed to an electric field
    • DNA moves because repelled by negative electrode end of electrophoresis
    • smaller DNA fragments move faster, different size fragments get separated from each other
    • only uses introns
  • polymerase chain reaction:: produces many copies of DNA without using a cell
    • DNA is treated with primers on either side of desired gene
    • put into a thermocycler with nucleotide and Taq polymerase
  • plasmid based transformation:: exogenous/artificial DNA can be placed in a host cell to carry out desired functions
  • microarray:: denatured DNA is placed in a gel electrophoresis, single stranded DNA with desired sequence is radioactively tagged and place in gel, if it finds a match the glowing probe and attached DNA will be visible.