Outline a basic technique used for gene transfer involving plasmids, a host cell (bacterium, yeast or other cell), restriction enzymes (endonucleases) and DNA ligase.

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The human gene that codes for insulin can be inserted into a plasmid and then this plasmid can be inserted into a host cell such as a bacterium. The bacterium can then synthesis insulin which can be collected and used by diabetics. This is done as follows.

The messenger RNA which codes for insulin is extracted from a human pancreatic cell which produces insulin using restriction enzymes. DNA copies are then made from this messenger RNA by using the enzyme reverse transcriptase and these DNA copies are then given extra guanine nucleotides to the end of the gene to create sticky ends.

At the same time, a plasmid is removed from a bacterial cell (plasmids are small, circular DNA molecules that can exist and replicate autonomously). A selected plasmid is cut using restriction enzymes endonuclease, which cut the DNA at specific base sequences. Then extra cytosine nucleotides are added to create sticky ends.

Once we have both the plasmid and the gene ready, these are mixed together. The two will link by complementary base pairing (between cytosine and guanine) and then DNA ligase is used to make the sugar phosphate bonds.

The plasmids with the human insulin gene (called recombinant plasmids) can then be mixed with host cells such as bacterium. The bacterium will take in the plasmid and start producing insulin, which can then be collected and purified.

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