Lecture 6-LC710 2nd hr

Lecture 6-LC710 2nd hr - Maxim-Gilbert Sequencing YNPV Logo...

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Plasmid (BAC) Phagemid Cosmid Lambda (PAC) Lots of tools for cloning:
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Needs to be at least 40kb to grow, but A left and right arm can be made smaller… See next slide
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Intron and Exon in Eukaryotic Cells mRNA DNA 5’ 3’ cap poly A tail exon exon exon intron intron mature mRNA Processing Transcription Splicing promotor 5’ 3’ Take place in nucleus start codon stop codon To cytoplasm Intron deleted
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Eukaryotes Splice Signals AG/GT AG/GT CAG/NT CAG/NT exon 1 intron 1 exon 2 exon 1 intron 1 exon 2
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cDNA Is Reverse Transcribed from mRNA mature mRNA poly A tail 5’ 3’ TTTT Reverse transcription CCC 3’ 5’ 3’ 5’ 3’ GGG DNA polymerase RNA hydrolysis 5’ 3’ 5’ Baltimore, Dulbecco, Temin (1975)
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cDNA Library mRNA cDNA Reverse transcription Genes in expression Complete gene Vector: Plasmid
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Libraries: cDNA cDNA library has sequences only from genes transcribed into mRNA (~3% of genome)
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MNGTEGPNFYVPFSNKTGVVRSPFEAPQYYLAEPWQFSMLAAYMFLLIML
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Unformatted text preview: GFPINFLTLYVTVQHKKLRTPLNYILLNLAVADLFMVFGGFTTTLYTSLH GYFVFGPTGCNLEGFFATLGGEIALWSLVVLAIERYVVVCKPMSNFRFGE NHAIMGVAFTWVMALACAAPPLVGWSRYIPEGMQCSCGIDYYTPHEETNN ESFVIYMFVVHFIIPLIVIFFCYGQLVFTVKEAAAQQQESATTQKAEKEV TRMVIIMVIAFLICWLPYAGVAFYIFTHQGSDFGPIFMTIPAFFAKTSAV YNPVIYIMMNKQFRNCMVTTLCCGKNPLGDDEASTTVSKTETSQVAPA* Maxim-Gilbert Sequencing MNGTEGPNFYVPFSNKTGVVRSPFEAPQYYLAEPWQFSMLAAYMFLLIML GFPINFLTLYVTVQHKKLRTPLNYILLNLAVADLFMVFGGFTTTLYTSLH GYFVFGPTGCNLEGFFATLGGEIALWSLVVLAIERYVVVCKPMSNFRFGE NHAIMGVAFTWVMALACAAPPLVGWSRYIPEGMQCSCGIDYYTPHEETNN ESFVIYMFVVHFIIPLIVIFFCYGQLVFTVKEAAAQQQESATTQKAEKEV TRMVIIMVIAFLICWLPYAGVAFYIFTHQGSDFGPIFMTIPAFFAKTSAV YNPV IYIMMNKQFRNCMVTTLCCGKNPLGDDEASTTVSKTETSQVAPA* Logo for a) Splice acceptor b) Splice Donor c) Initiator Met AG/GT AG/GT CAG/NT CAG/NT exon 1 intron 1 exon 2 exon 1 intron 1 exon 2...
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Lecture 6-LC710 2nd hr - Maxim-Gilbert Sequencing YNPV Logo...

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