Biotechnology

• genetic engineering refers to the techniques used to alter the chemical nature of genetic nature of genetic material and their introduction into another organism to change the phenotypic characters of that organism
• Paul Berg is known as the father of genetic engineering
• bioprocess engineering are used to maintain microbial contamination free conditions in chemical engineering processes
• A plasmid in an autonomously replicating extra-chromosomal circular DNA in some bacteria
• exonucleases remove nucleotides from the ends of the DNA in one strand of duplex
• endonucleases make cuts at specific position within DNA
• palindromes are the group of letters that form the same words when read both forward and backward
• DNA ligase is an enzyme that repais broken DNA by joining two nucleotides
• the DNA molecule that can carry a foreign DNA segment and replicate inside the host cell is called vector
• plasmids are small, autonomously replicating usually circular, extrachromosomal double stranded DNA molecule that occurs in many bacteria and some yeasts
• Alkaline phosphate removes the phosphate groups from the 5’ end of DNA molecule, leavening a free 5’ hydroxyl group
• bacteriophages are viruses infecting bacteria
• Transformation is a procedure through which a piece of DNA is introduced into the host bacterium
• cloning sites are generally required to ling the foreign or alien DNA with the vector
• DNA ligands are enzymes that help in repairing of broken DNA by joining two nucleotides
• alkaline phosphate is an enzyme that removes the phosphate group from the 5’ end of DNA molecule leaving a free 5‘ hydroxyl group
• origin of replication is a sequence from where replication starts
• Biolistics is another method suitable for introduction of DNA into plants
• RNA can be removed by treating it with ribonuclease
• preteins can be removed by its treatment with protease
• the material used in gel electrophoresis is agarose which is a natural polymer made of sea weeds
• the separated DNA can be visualized after only staining the DNA with Ethidium Bromide which is followed by UV radiation
• these separate strands of DNA are extracted from gel piece by a process known as elution
• PCR - polymerase chain reaction
• primers are chemically synthesized oligonucleotides that are complementary to the regions of DNA
• DNA template is the double stranded DNA that needs to be amplified
• The double stranded DNA is denatured by using high temperature
• annealing - two sets of oligonucleotide primers are annealed to the separated single strands
• the ampicillin resistance gene in this case is called selectable marker
• Any protein encoding gene expressed in a heterologous host, it is called recombinant protein.
• A bioreactor provides the optimal conditions for achieving the desired product by providing optimum growth conditions like temperatur, pH, substrate, salts, vitamins and oxygen
• components of a bioreactor

  • an agitator system
  • an oxygen delivery system
  • a foam control system
  • a temperature control system
  • pH control system
  • a sampling port to withdraw culture periodically
• a simple stirred tank bioreactor is usually cylindrical or with a curved base to facilitate even mixing of reactor components
• the sparged stirred tank bioreactor also facilitates the mixing of components and ensures oxygen availability throughout the bioreactor
• downstream processing

  • Separation of product from reactor
  • purification of the product
  • the product has to be manufactured, such formulations has to undergo clinical trials
  • strict quality control testing for each product is also required
• steps of recombinant DNA technology

  • Isolation of the genetic material
  • fragmentation of DNA by a restriction endonuclease
  • separation and isolation of a desired DNA fragment
  • amplification of gene of interest using PCR
  • ligation of the DNA fragment into vector
  • insertion of recombinant DNA into the host
  • obtaining or culturing foreign product
• Purified DNA will precipitate out after the addition of chilled ethanol
• collection of fine threads in the suspension which can be removed by spooling
• The longer the DNA fragment, the slower it moves through the gel
• Agarose gels separate DNA fragments based on their length