Genome projects such as DNA sequencing have read the genomes of a wide range of organisms including humans. The presence of non-coding DNA and regulatory genes menas knowledge of genome of more complex organisms cannot be easily translated into their proteome.
DNA fragments that we desire can be isolated or created by 3 ways: Using Reverse Transcriptase, Using Restriction Endonuclease and Using a Gene Machine.Download PDF (2 pages)
DNA fragments isolated from either reverse transcriptase or restriction endonucleases can then be cloned either using in vitro or in vivo cloning. In vivo cloning (inside organism) occurs by binding the sticky ends of a plasmid and the DNA fragment together and then growing the bacterial plasmid. The fragment and the plasmid are joined uisng DNA ligase.
In vitro cloning involves the polymerase chain reaction (PCR). First, the DNA mixture is heated to break hydrogen bonds and then cooled to allow primers to anneal to the strand. Te mixture is then heated again to allow free DNA nucloetides to bind to each template strand so DNA polymerase can work. Finally, two new copies of the fragment DNA are formed and 1 cycle of PCR is complete.
Recombinant DNA can be made by introducing foreign DNA into microorganisms by inserting it into a plasmid vector and using DNA ligase. The produced recombinant DNA then can be cloned by in vitro or in vivo cloning.
Gene probes contain a short sequence of DNA that is complementary to the gene you are looking for. If the allele is present, DNA will hybridise to it and can be detected with a label such as UV light or use of antibiotic resistance gene.
Genetic fingerprinting makes use of VNTR comparison using electrophoresis. DNA band patterns are produced on an electrophoresis gel depending on the number of VNTRs in each DNA.