Genomes are mapped to identify which parts of the genome that they come from. Information that is already known is used, such as the location of microsatellites.
Samples of the genome are mechanically sheared into smaller sections of around 100,000 base pairs.
These sections are placed in separate Bacterial Artificial Chromosomes (BACs) and transferred to E. coli cells. As the cells are grown in culture, many copies of the sections are produced - referred to as Clone Libraries.
Cells containing specific BACs are taken and cultured. The DNA is extracted from the cells and restriction enzymes are used to cut it into smaller fragments. The use of different restriction enzymes on a number of samples gives different fragment types.
The fragments are separated using electrophoresis.
The many copies of the fragments are put in a reaction mixture containing DNA polymerase, free DNA nucleotides and primers, with some of the nucleotides containing a flurescent markers. The primer anneals to the 3' end of the template strand, allowing DNA polymerase to attach and add free nucleotides.
If a modified nucleotide is affed, the polymerase enzyme is thrown off and the reaction stops on that template strand.
As the reaction proceeds, many molecules of DNA are made. The fragments generally differ in size, as different numbers of nucleotides will have been added before a modified nucleotide has been added.
As the strand runs through a machine, a laser reads the colour sequence, based on the length on the strands. The sequence of colours, and therefore the sequence of bases, can be displayed.
The identification of genes for proteins found in all or many living organisms gives clues to the relative importance of such genes to life.
Comparing the DNA of different species shows evolutionary relationships. Modelling the effects of changing DNA can be carried out. Comparing genomes from pathogentic and similar but non-pathogenic organisms can be used to identify the genes or base-pair sequences that are more important in causing the disease, so more effective drugs can be developed. The DNA of individuals can be analysed to reveal the presence of alleles associated with particular diseases.
Recombinant DNA: A section of DNA, often in the form of a plasmid, which is formed by joining DNA sections from two different sources.
Genetic engineering involves the extraction of genes from one organism, or the manufacture of genes, in order to place them into another organism (often of a different species) such that the receiving organism expresses the gene product. The required gene is obtained. A copy of the gene is placed in a vector. The vector carries the gene to the recipient cell. The recipient expresses the gene through protein synthesis.
Sections of DNA containing a desired gene can be extracted from a donor organism using restriction enzymes. Use a restriction enzyme (endonuclease) to cut out the gene coding for the protein required or to fragment (digest) the DNA and use a gene probe. This produces sticky ends.
Or - obtain mRNA and then use reverse…