BIOL 200 Lecture Notes - Lecture 23: Autoradiograph, Amniote, Recombinant Dna

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Cdna library is a complete or partially complete collection of coding sequences, exclusively; no transcripts. Library screening to retrieve a particular sequence in millions of bacteria, use hybridization. Place bacteria on agar plates, pick clones that are of interest; make a replica by placing nitrocellulose filter on top to get bacteria on nitrocellulose incubate filter in alkaline solution to denature dna. Hybridize dna with labeled probe; wash away labeled dna that does not hybridize to dna bound to filter; perform autoradiography. Go back to original plate to pick out the gene. Expression of recombinant proteins recombinant proteins can be expressed using bacteria. Start with cdna (coding sequence), which is taken from mrna. A promoter is needed to start transcription of the sequence. Place recombinant vector containing cdna and promoter and viral origin of replicationinto cultured cells by lipid treatment + electroporation (poke transient hole in cell membrane)

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