6.3.6(Genetic engineering)

Describe the necessary stages that are necessary for genetic engineering

• The required gene is isolated

• A copy of the gene is placed inside of a vector

• Transform host cells with the vector

• Identify the host cells that have taken up the gene using gene markers

• Grow or clone the host cells

Describe what is meant by a recombinant plasmid

• It is a plasmid which carries a gene from a different organism

Describe what is meant by a transgenic bacterium

• It is a a bacterial cell which contains the recombinant plasmid

what are restriction enzymes described as

• They are palindromic

• They are many types, each recognising specific short DNA sequences

Describe how restriction enzymes are useful

• They recognise specific sequences of DNA

• They cut the DNA sequences, either as a staggered cut or as blunt end cuts

Describe the process of isolating the gene

• Use restriction enzymes to cut gene out of cell they cut using a staggered mechanism to leave sticky ends which allows for exposed bases which allows for easy binding to the plasmid

Describe the process of placing the gene into a vector

• Plasmids from bacteria or other organisms can be used as a vector

• The same restriction enzyme can be used to cut specific site of the plasmid

• The cut plasmid has exposed sticky ends

• Complementary nucleotides stick to the sticky ends of the plasmid

• The gene and the plasmid anneal using DNA ligase enzyme which forms phosphodiester bonds between the gene and the plasmid

• The plasmid is then inserted into a host cell

Describe the method of transformation

• Electroporation is used

  • This is where a high voltage pulse is applied to the cell to disturb the membrane

  • This increases the permeability of the cell membrane which allowed the plasmid to enter into the cell

Describe what occurs after transformation of a transgenic bacterium

• The bacteria that have been transformed are used to make pure cultures

• These cultures are grown in large fermenters

• They divide by binary fission rapidly to reproduce to form genetically identical daughter cells

Describe the process of genetic engineering

• Isolate the gene of interest and cut from donor using a restriction enzyme

• Use the same restriction enzyme to cut a plasmid - A vector - then insert the gene

• Transform host cells with the vector using electroporation

• Use gene markers to identify that the host cells that have taken up the gene

• Grow and clone the host cell

Describe the use of gene markers in genetic engineering

• Gene markers are used to identify whether a gene has been taken up by bacterial cells

• A fluorescent protein that is easily seen or an antibiotic resistant bacteria is used as a marker

Explain why transformation must occur during genetic engeineering

• It occurs because DNA does not easily cross the host cell plasma membrane

Describe the advantages of genetic engineering

• Large fragments of DNA can be clones

• Produces transformed bacteria that can used to produce large quantities of gene products

• Relatively cheap method

• Almost no risk of contamination

• Can produce mRNA and protein as well as DNA because it is done in a living cell

Describe the disadvantages of invivolobing

• The DNA fragment has to be isolated from the host DNA or cell components

• The process takes a long time and requires large fermenters