Lecture 15 new Biol302 Spring 2011

Lecture 15 new Biol302 Spring 2011 - Blue/White Selection...

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Blue/White Selection
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Alpha complementation Trick alpha omega
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Blue/White Selection
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two proteins/same mRNA Bicistronic strategy:Labels Neurons β gal β gal
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two proteins/same mRNA Bicistronic strategy uses IRES β gal β gal IRES: Internal Ribosome Entry Site
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M71-IRES-Tau lacZ M71 M71
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add Fe +2 Fe
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add FastREDViolet FRV
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M71 -IRES- LacZ ( FRV-Xgal ) M72 -IRES- GFP ( Green Fluorescent Protein )
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Foundations of a replicative organism
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DNA Discovery by Friedrich Miescher (Swiss, 1844-1895) He discovered a substance containing both phosphorus and nitrogen, made up of molecules that were apparently very large, in the nuclei of white blood cells Named the substance nuclein because it seemed to come from cell nuclei. In 1874 when Miescher separated it into a protein and an acid molecule. It is now known as deoxyribonucleic acid (DNA)
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He worked with Albrecht Kossel and Emil Fischer, the nucleic acid and protein experts at the turn of the 20 th . century Phoebus Levene
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He conducted experiments that in 1931 suggested that the four components of DNA occur in approximately equal ratios He suggested the possibly that DNA was made of a repeating tetramer If so, the implication was that the structure of DNA was too simple and too regular to contribute to genetic variation: attention thereafter focused on protein as the probable hereditary substance
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Not only did Levene identify the components of DNA, he also showed that the components were linked together in the order phosphate-sugar-base to form units. He called each of these units a nucleotide, and stated that the DNA molecule consisted of a string of nucleotide units linked together through the phosphate groups, which are the 'backbone' of the molecule. Scientist thought that Proteins (made from 20 aa) were Needed to encode life, not the 4(?) forms of Nucleotides
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The Tetranucleotide Hypothesis
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What is Life? that planted the idea for searching “the secret of life”
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Lecture 15 new Biol302 Spring 2011 - Blue/White Selection...

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