33_Lieb_blotting_RFLPs_1

33_Lieb_blotting_RFLPs_1 - Recombinant DNA III: Blotting...

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Unformatted text preview: Recombinant DNA III: Blotting & RFLPs J. Lieb April 14, 2008 1 2 2 2 2 2 2 2 2 2 2 2 1 2 1 2 1 2 1 2 1 2 d d D d D d D d D d D d D d d d d d d d A Typical Expression Vector Last Lecture Vectors cDNA and Libraries grows into colony containing fragment 1 grows into colony containing fragment 2 Genomic DNA libraries partial digestion restriction map of genomic DNA (each line represents one restriction site) some fragments produced by partial digestion size selection fragments between 10 and 20 kb Creating a genomic DNA library So, now weve got our gene (or at least part of it) into the right vector How can I find my gene of interest in a library? Can I find other clones that contain the same gene? Are there are rearrangements in that gene in genomic DNA? Where and when is RNA from that gene produced? Where and when is Protein from that gene produced? Screening Colonies by Hybridization Some Probe sources: cDNA from a tissue expressing the gene highly Homologous gene Sequence derived from the protein product of gene of interest So, now weve got our gene (or at least part of it) into the right vector How can I find my gene in a library? Can I find other clones that contain the same gene?...
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33_Lieb_blotting_RFLPs_1 - Recombinant DNA III: Blotting...

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