plasmids

Examples of plasmids in the following topics:

• Types of Plasmids and Their Biological Significance

  • Plasmids used in genetic engineering are called vectors .
  • Many plasmids are commercially available for such uses.
  • Only bacteria that take up copies of the plasmid survive, since the plasmid makes them resistant.
  • Another way to classify plasmids is by function.
  • Fertility F-plasmids, which contain tra genes.

• Introduction to Plasmids

  • Plasmid sizes vary from 1 to over 1,000 kbp.
  • The term plasmid was first introduced by the American molecular biologist Joshua Lederberg in 1952.
  • Plasmids are considered replicons .They can be found in all three major domains: Archaea, Bacteria, and Eukarya.
  • Similar to viruses, plasmids are not considered by some to be a form of life.
  • The upper half of the image shows a bacterium with its chromosomal DNA and plasmids dividing into two identical bacteria, each with their chromosomal DNA and plasmids.

• Agrobacterium and Crown Gall Disease

  • Argobacterium causes Crown Gall Disease by transferring a DNA plasmid to the host plant, causing the host to make nutrients for it.
  • A. tumefaciens can transfer part of its DNA to the host plant, through a plasmid - a bacterial DNA molecule that is independent of a chromosome.
  • For example, plasmids can confer on bacteria the ability to fix nitrogen, or to resist antibiotic compounds.
  • Typically bacteria transfer plasmids through conjugation: a donor bacteria creates a tube called a pilus that penetrates the cell wall of the recipient bacteria and the plasmid DNA passes through the tube.
  • The other bacteria either integrates the plasmid into its chromosomes, or it remains free-floating in the cytoplasm.

• Vectors for Genomic Cloning and Sequencing

  • The four major types of vectors are plasmids, viral vectors, cosmids, and artificial chromosomes.
  • Plasmid vectors minimalistically consist of an origin of replication that allows for semi-independent replication of the plasmid in the host and also the transgene insert.
  • In the case of plasmids utilized as transcription vectors, incubating bacteria with plasmids generates hundreds or thousands of copies of the vector within the bacteria in hours.
  • Plasmids may be conjugative / transmissible and non-conjugative.
  • Conjugative vectors mediate DNA transfer through conjugation and therefore spread rapidly among the bacterial cells of a population, such as the F plasmid, as well as many R and some col plasmids.

• Plasmids and Lysogeny

  • Plasmids are DNA molecules that are capable of replicating independently from the chromosomal DNA.
  • Plasmids are often characterized by their circular appearance and double-strands; they also vary in size and number.
  • 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.
  • Specifically, the exchange of plasmid DNA falls under transformation.

• Plasmids as Cloning Vectors

  • Plasmid vectors minimally consist of the transgene insert and an origin of replication, which allows for semi-independent replication of the plasmid in the host.
  • Plasmids may be conjugative/transmissible or non-conjugative.
  • Nonconjugative plasmids do not mediate DNA through conjugation.
  • The pGEX-3x plasmid is a popular cloning vector.
  • The various elements of the plasmid are labelled.

• Shuttle Vectors and Expression Vectors

  • An expression vector is generally a plasmid that is used to introduce a specific gene into a target cell.
  • An expression vector, otherwise known as an expression construct, is generally a plasmid that is used to introduce a specific gene into a target cell .
  • The plasmid is frequently engineered to contain regulatory sequences that act as enhancer and promoter regions and lead to efficient transcription of the gene carried on the expression vector.
  • Cloning vectors, which are very similar to expression vectors, involve the same process of introducing a new gene into a plasmid, but the plasmid is then added into bacteria for replication purposes.
  • The pGEX-3x plasmid is a popular cloning vector.

• Genetic Analysis

  • Bacterial chromosomes contain double stranded molecules of DNA arranged in a circular form called plasmids .
  • Plasmids are located in the cytoplasm of bacteria, are capable of autonomous replication, and transfer genes from parent cell to daughter cell.
  • Plasmid profiling using molecular, biochemical, and microbial techniques is essential to understanding the mechanism of pathogenicity and to fuel genetic engineering.

• Hosts for Cloning Vectors

  • A very large number of host organisms and molecular cloning vectors are in use, but the great majority of molecular cloning experiments begin with a laboratory strain of the bacterium E. coli (Escherichia coli) and a plasmid cloning vector.
  • E. coli and plasmid vectors are in common use because they are technically sophisticated, versatile, widely available, and offer rapid growth of recombinant organisms with minimal equipment.
  • In practice, however, specialized molecular cloning experiments usually begin with cloning into a bacterial plasmid, followed by subcloning into a specialized vector.

• Mammalian Gene Expression in Bacteria

  • A DNA sequence for a protein of interest could be cloned or subcloned into a high copy-number plasmid containing the lac promoter, which is then transformed into the bacterium Escherichia coli.
  • The genetically engineered expression system contains the appropriate DNA sequence for the gene of choice which is engineered into a plasmid that is introduced into a bacteria host.
  • In a genetically engineered system, this entire process of gene expression may be induced depending on the plasmid used.
  • With E. coli, DNA is normally introduced in a plasmid expression vector.