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Lecture 37-Gene Technology

Lecture 37-Gene Technology - Lecture 37 Gene Technology...

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Lecture 37- Gene Technology Transgenesis -Moving DNA from one cell to another -Requires: a) Isolating the DNA of interest b) A vector: a system of getting DNA into a host (often a plasmid or a virus) c) Mechanism of transfer d) Selectable “marker” DNA of Interest -DNA to be transferred may be: a) Isolated (cut) directly from a genome b) Created by reverse transcription of mRNA called “cDNA” (complimentary DNA) c) Synthesised in a lab Vectors -Modified or artificial plasmids, viruses or chromosomes -“Sticky ends” produced by digestion with restriction endonucleases facilitate creation of “recombinant DNA” Mechanisms of Transfer -Moving across host cell membrane/wall: a) Transformation of “naked” DNA (electroporation: cells in an electric field with DNA, pushes DNA into cell) b) Transfection (inside viral capsid) c) Liposomes (artificial membrane) d) Microinjection e) Biolistics (gene gun) Selectable Marker -Vectors carry genes coding for a phenotype that identifies host cells carrying new DNA: a) Antibiotic resistance b) Reporter genes: fluorescence or colour or light (makes it obvious a vector gene has been taken by cell) Gene Cloning 1) Isolate genomic DNA containing gene of interest from cells and cut the DNA into fragments 2) Cut a circular bacterial plasmid to make it linear 3) Insert the genomic DNA fragments into the plasmid to make recombinant DNA molecules. Recombinant DNA is from two different sources joined together. When using a circular plasmid, the inserted fragment of DNA makes the plasmid the recombinant.
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