recombinant DNA technology

Examples of recombinant DNA technology in the following topics:

• Biochemical Products of Recombinant DNA Technology

  • Many practical applications of recombinant DNA are found in human and veterinary medicine, in agriculture, and in bioengineering.
  • Recombinant DNA technology is the latest biochemical analysis that came about to satisfy the need for specific DNA segments.
  • Recombinant DNA technology engineers microbial cells for producing foreign proteins, and its success solely depends on the precise reading of equivalent genes made with the help of bacterial cell machinery.
  • Recombinant DNA technology, apart from being an important tool of scientific research, has also played a vital role in the diagnosis and treatment of various diseases, especially those belonging to genetic disorders.
  • Some of the recent advances made possible by recombinant DNA technology are:

• Recombinant DNA Technology

  • Recombinant DNA technology also referred to as molecular cloning is similar to polymerase chain reaction (PCR) in that it permits the replication of a specific DNA sequence.
  • To improve the ratio of recombinant to non-recombinant organisms, the cleaved vector may be treated with an enzyme (alkaline phosphatase) that dephosphorylates the vector ends.
  • The creation of recombinant DNA is in many ways the simplest step of the molecular cloning process.
  • Insertion of the foreign DNA into the beta-galactosidase coding sequence disables the function of the enzyme, so that colonies containing recombinant plasmids remain colorless (white).
  • Therefore, recombinant clones are easily identified .

• Mammalian Proteins and Products

  • Recombinant DNA technology not only allows therapeutic proteins to be produced on a large scale but using the same methodology protein molecules may be purposefully engineered.
  • Bacterial expression systems, due to their simplicity, are often not able to produce a recombinant human protein identical to the naturally occurring wild type.
  • Recombinant clotting factors have eliminated this problem.

• Large-Scale Fermentations

  • Fermentation is also utilized in the mass production of various recombinant products.
  • These recombinant products include numerous pharmaceuticals such as insulin and hepatitis B vaccine.
  • The mass production of insulin is performed by utilizing both recombinant DNA technology and fermentation processes.
  • An additional recombinant product that utilizes the fermentation process to be produced is the hepatitis B vaccine.
  • The creation of this vaccine utilizes both recombinant DNA technology and fermentation.

• Plasmids and Lysogeny

  • Plasmids are DNA molecules that are capable of replicating independently from the chromosomal DNA.
  • 'Naked DNA' refers to a specific type of DNA which does not encode for genes promoting the transfer of genetic material to a new host.
  • In the field of molecular biology, plasmid DNA is often referred to as 'vectors' due to their ability to transfer DNA between organisms.
  • The use of plasmid DNA in molecular biology is considered to be recombinant DNA technology.
  • The advantages of plasmid DNA transfer allow for survival advantages.

• Generalized Recombination and RecA

  • In homologous recombination, a type of genetic recombination, nucleotide sequences are exchanged between two similar molecules of DNA.
  • Homologous recombination is a type of genetic recombination in which nucleotide sequences are exchanged between two similar or identical molecules of DNA.
  • Homologous recombination is a major DNA repair process in bacteria.
  • Double-strand DNA breaks in bacteria are repaired by the RecBCD pathway of homologous recombination .
  • The RecBCD pathway is the main recombination pathway used in bacteria to repair double-strand breaks in DNA.

• Selection

  • DNA recombination has been used to create gene replacements, deletions, insertions, inversions.
  • Subsequently, these fragments are then combined with vector DNA to generate recombinant DNA molecules.
  • This will generate a population of organisms in which recombinant DNA molecules are replicated along with the host DNA.
  • Thus, both the resulting bacterial population, and the recombinant DNA molecule, are commonly referred to as "clones".
  • Strictly speaking, recombinant DNA refers to DNA molecules, while molecular cloning refers to the experimental methods used to assemble them.

• Obtaining DNA

  • Molecular cloning is a set of experimental methods in molecular biology that are used to assemble recombinant DNA molecules and to direct their replication within host organisms.
  • Molecular cloning generally uses DNA sequences from two different organisms: the species that is the source of the DNA to be cloned, and the species that will serve as the living host for replication of the recombinant DNA.
  • The creation of recombinant DNA is in many ways the simplest step of the molecular cloning process.
  • Whichever method is used, the introduction of recombinant DNA into the chosen host organism is usually a low efficiency process; that is, only a small fraction of the cells will actually take up DNA.
  • Explain the methods of obtaining DNA for molecular cloning experiments and the process of creating a recombinant DNA molecule

• Bacterial Transduction

  • There are generally three types of recombination events that can lead to this incorporation of bacterial DNA into the viral DNA, leading to two modes of recombination.
  • This may occur in two main ways, recombination and headful packaging.
  • This bacterial material may become recombined into another bacterium upon infection.
  • When the new DNA is inserted into this recipient cell it can fall to one of three fates: the DNA will be absorbed by the cell and be recycled for spare parts; if the DNA was originally a plasmid, it will recirculate inside the new cell and become a plasmid again; if the new DNA matches with a homologous region of the recipient cell's chromosome, it will exchange DNA material similar to the actions in conjugation.
  • This type of recombination is random and the amount recombined depends on the size of the virus being used.

• Molecular Products from Microbes

  • The isolation of approximately 3000 restriction enzymes has allowed molecular biologists to utilize them in processes such as cloning and the production of recombinant DNA .
  • DNA ligase plays a key role in molecular biology processes due to its ability to insert DNA fragments into plasmids.
  • The process of DNA ligation is defined as the ability of DNA ligase to covalently link, or ligate, fragments of DNA together.
  • In molecular biology -- specifically, during the process of developing recombinant DNA -- DNA ligase can be used to ligate a fragment of DNA into a plasmid vector .
  • The most commonly used DNA ligase is derived from the T4 bacteriophage and is referred to as T4 DNA ligase.